WEBVTT 00:00.000 --> 00:09.560 Through where I think I'm at, now I'm going to give you my hypothesis in a nutshell and 00:09.560 --> 00:15.920 then I'm going to drink a beer and probably light a fire and see if I can read some comments 00:15.920 --> 00:18.520 before I go to bed. 00:18.520 --> 00:23.880 My current hypothesis is this, this likely originated from a laboratory source. 00:23.880 --> 00:29.000 Number one, laboratory leaks happen and you can look up many different articles including 00:29.000 --> 00:36.060 a few from Lynn Klotz who will show you definitively through history how leaks have happened through 00:36.060 --> 00:43.080 unintended infection, through waste products, lots of different ways and lots of different 00:43.080 --> 00:45.440 places it has happened. 00:45.440 --> 00:50.040 The second reason I believe this is true is because of the circumstantial bullseye, the 00:50.040 --> 00:56.160 idea that this broke out in Wuhan where the research on coronavirus was happening where 00:56.160 --> 01:01.360 the foremost groups were working on the foremost dangerous viruses and even though you want 01:01.360 --> 01:06.160 to say and people will tell you that this was BSL Level 4, most of the coronavirus work 01:06.160 --> 01:12.600 that is published in the literature is done in BSL Level 2 or BSL Level 3, even BSL Level 01:12.600 --> 01:19.960 3 is not a very strict biosafety level, I work in a BSL 3 level lab with my mice. 01:19.960 --> 01:24.600 And finally, the entangled financial interests I think are also very important to acknowledge 01:24.600 --> 01:28.280 here the amount of money that was given away to private equity, the amount of money 01:28.280 --> 01:35.040 that has been given away without context or identification to millions of corporations 01:35.040 --> 01:40.000 in America and the amount of money that is being given away through the Warp Speed Act 01:40.000 --> 01:47.400 that's allowing big pharma companies to make products that are guaranteed to be purchased 01:47.400 --> 01:52.560 or to be supported by government funding under the pretense that we need to bring as 01:52.600 --> 01:56.440 much of this to bear as possible. 01:56.440 --> 01:59.600 One of the things that you have to realize here, and I hadn't made this point yet but 01:59.600 --> 02:07.400 I know I'm glad I remembered, there is a federal law, the Defense Production Act, which allows 02:07.400 --> 02:14.240 the government to tell companies and corporations that they need to produce certain things at 02:14.240 --> 02:20.120 cost because it is for the benefit of all of the American people, for the benefit of 02:20.120 --> 02:22.280 America as a whole. 02:22.280 --> 02:26.560 This was an act that was designed to change the production of companies during World War 02:26.560 --> 02:34.200 One or World War Two and we could easily use this to direct pharmaceutical companies to 02:34.200 --> 02:40.800 develop specific things like, for example, produce hydrochloroquine or produce a vaccine 02:40.800 --> 02:46.880 of a particular design or a particular type and we could limit and control the amount 02:46.880 --> 02:52.800 of profit that was made so that nothing was, no one became rich from it. 02:52.800 --> 02:55.840 We could do the same thing with coronavirus testing. 02:55.840 --> 03:00.960 We could be using that act to make sure that no one is profiting from the testing, from 03:00.960 --> 03:06.400 the antibody testing and from the PCR testing, which I'm going to do another whole talk about 03:06.400 --> 03:12.640 that but PCR as a test is absurd right now and we know that because this test is not 03:12.640 --> 03:18.920 specific for SARS-CoV-2 but is only specific for SARS viruses in general and that means 03:18.920 --> 03:23.040 that we could, as I said before, be testing for the endemic version of the descendants 03:23.040 --> 03:27.680 of the original SARS, which are almost guaranteed to have gone through at least half of our 03:27.680 --> 03:32.120 population in the last decade and a half. 03:32.120 --> 03:36.200 So SARS-1 and its descendants have gone endemic, that's what I was just saying. 03:36.200 --> 03:42.560 The current PCR test is not specific for SARS-CoV-2 and that is known because it's the German 03:42.600 --> 03:46.480 company that originally produced it, knows that they've chosen a short sequence that 03:46.480 --> 03:47.480 they think is relevant. 03:47.480 --> 03:53.320 It is a short sequence that just comes from one of the open reading frame one proteins. 03:53.320 --> 03:58.520 We are being duped by a non-specific test and we would have gotten these numbers or 03:58.520 --> 04:03.880 numbers very similar to them with or without this pandemic because we've never looked for 04:03.880 --> 04:07.560 SARS viruses as they are spread through our population. 04:07.560 --> 04:09.600 We've just never looked. 04:09.600 --> 04:12.520 Nobody sampled for it until this year. 04:12.520 --> 04:19.000 It's a huge point to make because the SARS virus originally appeared in 2003 and disappeared 04:19.000 --> 04:25.120 in 2004 but it didn't really disappear, it just became not a problem. 04:25.120 --> 04:32.440 Descendants of that virus of various virulence and infectivity have circulated in the time 04:32.440 --> 04:40.440 since they didn't disappear and this test is not specific for any one of them and no one 04:40.440 --> 04:45.920 has ever done a study about how many descendants there are, how many people are infected, 04:45.920 --> 04:51.000 how many animals have been infected by it, etc. 04:51.000 --> 04:59.280 So that's a pretty interesting place to be in July of 2020. 04:59.280 --> 05:06.240 Thinking that there are descendants of the original SARS leak or leaks or zoonotic jumps 05:06.240 --> 05:12.240 which are building up an endemic signal in the background that could then be turned 05:12.240 --> 05:18.400 on in the flick of a switch, just kind of like Kevin McCurnan explained it in the video 05:18.400 --> 05:24.480 interview with me and a few other people where he said you could do it with HKU1. 05:24.480 --> 05:26.600 Just turn on the PCR and start monitoring it. 05:26.600 --> 05:32.360 You're going to see patterns of viruses moving through the population according to him. 05:32.360 --> 05:38.040 So if there are SARS descendants which could be used and misconstrued in that way, certainly 05:38.040 --> 05:41.520 that was a possibility that should have been considered from the very beginning of the 05:41.520 --> 05:44.520 pandemic and it was not. 05:44.520 --> 05:50.680 Certainly wasn't something that Kevin McCurnan brought up during his authorship of the paper 05:50.680 --> 05:57.560 that was let's say opposing the PCR test, opposing the PCR test by saying that some 05:57.560 --> 06:00.840 of the primers would make primer dimers. 06:00.840 --> 06:08.880 Never mind the whole pretense of being specific for a specific virus when the PCR test that 06:08.880 --> 06:15.520 was designed by the German lab was using sequences that they had found in Germany, sequences 06:15.520 --> 06:19.080 from German bats. 06:19.080 --> 06:26.280 I mean there was a lot of other reasons to lash out against the PCR test that was designed 06:26.280 --> 06:32.920 by Jorsten but primer dimers and this kind of thing was an odd sort of objection and 06:32.920 --> 06:38.080 if you look back and see who was on that paper, he will see that a number of early objectors 06:38.080 --> 06:41.440 were all roped in together. 06:41.440 --> 06:45.760 I think we're going to be coming back to that little gathering of quote unquote dissidents 06:45.760 --> 06:50.240 many many times in the next year or so. 06:50.240 --> 06:55.080 And so saying that we're being duped by a nonspecific PCR test is also really not specific 06:55.080 --> 07:01.080 enough even though right now this particular version of me was not completely queued in 07:01.080 --> 07:06.440 to the fact that this time that there could have been as many as 150 different variants 07:06.440 --> 07:14.640 of the test, 150 different variants of the test being utilized and being employed by maybe 07:14.640 --> 07:20.280 hundreds if not thousands of independent labs with various levels of quality control and 07:20.280 --> 07:23.120 certainly no oversight. 07:23.120 --> 07:27.520 And the crazy thing is as many of those small company labs that were doing a lot of the 07:27.520 --> 07:33.520 local testing especially in places where the testing was mandated, those are all gone. 07:33.520 --> 07:35.920 They don't exist anymore. 07:35.920 --> 07:39.800 Those companies took millions of dollars and they're gone. 07:39.800 --> 07:43.720 Their owners own houses and they're gone. 07:44.720 --> 07:50.360 And we're supposed to focus on the DNA fragments in the shot and forget about the fact that 07:50.360 --> 07:58.920 the whole the whole theater in the beginning to make worst case scenario real was wholly 07:58.920 --> 08:01.080 created out of out of fresh cloth. 08:01.080 --> 08:08.200 This is not sorry to start the show like this, but this is just the way it happened to be. 08:08.200 --> 08:12.880 I had this queued up for some reason. 08:12.880 --> 08:17.040 So the next thing that I want to say is our lack of biological knowledge and the general 08:17.040 --> 08:23.120 poor health of the of America, the American people is being used to create the crisis 08:23.120 --> 08:26.560 they need to divide and conquer us to ruin. 08:26.560 --> 08:27.560 Maybe America. 08:27.560 --> 08:28.560 I don't know. 08:28.560 --> 08:29.560 Crash the dollar. 08:29.560 --> 08:30.560 I don't know. 08:30.560 --> 08:34.480 Steel the rest of our are what limited treasury value we have left. 08:34.480 --> 08:36.720 I don't know. 08:36.720 --> 08:42.360 But I know for sure that they are combining our lack of biological knowledge and our general 08:42.360 --> 08:49.120 society's lack of good health and access to health care to create a crisis to usher 08:49.120 --> 08:54.640 in all kinds of changes that would otherwise never be necessary, and more importantly never 08:54.640 --> 08:56.840 be possible. 08:56.840 --> 09:01.200 Just like the Patriot acted 9-11. 09:01.200 --> 09:06.760 Highly profitable control mechanisms based on immunity are the near term goal. 09:06.760 --> 09:11.040 I believe this firmly now, there are too many people talking about it in the mainstream 09:11.040 --> 09:13.920 media hinting at it. 09:13.920 --> 09:18.600 Sound you hear is a single cicada, there are no crickets. 09:18.600 --> 09:23.000 Highly profitable control mechanisms, I'm talking about apps that you have to have and 09:23.000 --> 09:27.680 maybe even worse in order to cross state lines, in order to go to work, maybe in order to 09:27.680 --> 09:31.560 go to the store, you're going to have this happen. 09:31.560 --> 09:36.760 So there's going to be technology that needs to be distributed, there's going to be laws 09:36.760 --> 09:40.560 that are going to be need to be passed, there's going to be all kinds of products that need 09:40.560 --> 09:47.000 to be sold, and the whole idea is going to be to sell you on the idea that antibodies 09:47.000 --> 09:51.960 are the only evidence of immunity, and the only way to get antibodies is to get this 09:51.960 --> 09:57.840 vaccine, because a lot of people of course are not showing long lasting antibody levels, 09:57.840 --> 10:02.200 so even the people who've been infected, and that's what they're hyping right now is that 10:02.200 --> 10:06.760 you can get infected again, this is the narrative they're bringing to you so that they're going 10:06.760 --> 10:12.480 to tell you, the only way we're going to be sure is that if everybody gets vaccinated, 10:12.480 --> 10:20.360 and now we have billions of doses necessary, billions of doses backed by government money, 10:20.360 --> 10:28.320 backed by government mandate, controlled by apps, distributed yearly, we are talking 10:28.320 --> 10:33.560 about a very serious change in our healthcare system that will benefit the largest corporations 10:33.560 --> 10:38.400 and allow them to profit, because there are no controls on that profit right now, none 10:38.400 --> 10:42.560 at all, we have chosen to do nothing of the sort. 10:42.560 --> 10:47.320 That's why I'm sure this hypothesis is correct, the other reason why I'm sure this hypothesis 10:47.400 --> 10:51.920 is correct is, well I'm getting more sure that this hypothesis is correct, I don't want 10:51.920 --> 10:57.200 to sound like non-scientific here or non-objective, is because the guy that I first really got 10:57.200 --> 11:03.680 woken up by on the internet besides Dan of Harvard to the big house blog, is Wolfgang 11:03.680 --> 11:09.360 Wodach, and for some reason I never searched for this guy after I saw this German video 11:09.360 --> 11:14.520 that I had to get translated by a friend of mine, where he explained that it was likely 11:14.520 --> 11:19.000 a SARS virus or a SARS-descended virus that has been circulating viruses that have been 11:19.000 --> 11:28.000 circulating since the original SARS outbreak fizzled out in 2004 that we are testing for, 11:28.000 --> 11:34.840 and he only released that video in May or in April and it was only in German, and then 11:34.840 --> 11:35.840 I just never... 11:35.840 --> 11:40.320 I hope you see how significant it is that Wolfgang and I have now made contact in such 11:40.320 --> 11:42.360 a significant way. 11:42.360 --> 11:47.880 I hope you see what kind of forward progress is about to happen, don't be distracted by 11:47.880 --> 11:54.920 what's going on on Twitter and how people supposedly think that this story of the clones 11:54.920 --> 12:00.480 is an open and shut case now because there's effectively no swarm. 12:00.480 --> 12:06.300 He's been going around in circles on this idea for a very, very long time, and the reason 12:06.300 --> 12:11.240 why he's doing that is because all of RNA virology is dependent on it and no one can 12:11.240 --> 12:12.440 know that. 12:12.440 --> 12:21.840 If everybody understands that, that what they speak of as virology is a gross exaggeration 12:21.840 --> 12:28.800 of fidelity and it's a gross exaggeration of understanding, even the next strain database 12:28.800 --> 12:34.880 and phylogeny, even the Jed said database or whatever the hell it's called, all of these 12:34.880 --> 12:45.360 collections of genomes are presented to you as gold standard evidence of and proof 12:45.360 --> 12:49.360 of something for which they are not proof of. 12:49.360 --> 12:53.360 All we know for sure is that these people have managed to measure these sequences and 12:53.360 --> 12:58.580 a lot of these people believe these sequences are real, but we don't have any reason to 12:58.580 --> 13:04.360 believe that these sequences are somehow related to one another in a progression related to 13:04.360 --> 13:07.960 one another in an evolutionary way. 13:07.960 --> 13:13.280 We don't even know, no one is even talking about the possibility that these animals and 13:13.280 --> 13:20.600 ourselves would emit coronavirus like signals from our immune system or from the outside, 13:20.600 --> 13:24.320 from our lungs. 13:24.320 --> 13:30.600 No one's even talking about the possibility that these signals are being produced by the 13:30.600 --> 13:41.160 bacteria in our lungs or the bacteria in our airways, our GI tracks, wherever. 13:41.160 --> 13:46.600 No one's talking about the possibility that there's replication of RNA or DNA being carried 13:46.600 --> 13:51.400 by bacteriophages that's doing these signals, that's causing these signals, which really 13:51.400 --> 13:58.720 are just signals, genetic signals that are coincident with, well, genetic signals that 13:58.720 --> 14:03.240 are present and they're coincident with amplicons of PCR. 14:03.240 --> 14:11.920 It's very difficult at this size scale and at this sensitivity to definitively say that, 14:11.920 --> 14:18.520 well, this is definitely a something and we must always end up taking the word of somebody 14:18.520 --> 14:20.120 like Kevin McCurnan. 14:20.120 --> 14:21.280 They can't prove it to you. 14:21.280 --> 14:24.120 He's not going to ever teach you how this stuff works. 14:24.120 --> 14:27.760 He's always going to tell you that it's just too complicated for him to explain it 14:27.760 --> 14:31.280 to you, but I'm definitely wrong. 14:31.280 --> 14:38.120 When in reality, the idea that we're fighting about is really very simple and that's why 14:38.120 --> 14:43.600 it keeps spiraling out of control behind a block where we can't really have this discussion 14:43.600 --> 14:49.040 and why he keeps saying that we're not really, that he had this discussion a long time ago 14:49.040 --> 14:53.240 on email or he tried to do this on his sub stack before. 14:53.240 --> 14:57.160 I've addressed all the papers in his sub stack when they came out. 14:57.160 --> 15:02.120 I did a show about it several times in a row, which he completely ignored. 15:02.120 --> 15:03.120 And that's fine. 15:03.120 --> 15:04.120 He doesn't have to watch it. 15:04.120 --> 15:06.440 He calls my videos six hours Scooby-Doo videos. 15:06.440 --> 15:07.440 So that's fine. 15:07.440 --> 15:08.440 I get it. 15:08.440 --> 15:12.040 I should write a sub stack for him. 15:12.040 --> 15:13.880 We're going to figure this out, ladies and gentlemen. 15:13.880 --> 15:14.880 Don't worry. 15:14.880 --> 15:19.600 And when we figure it out the way that the ball will move forward, it will move in a very 15:19.600 --> 15:23.200 different way with a very different direction and a very different momentum. 15:23.200 --> 15:31.000 I assure you, I assure you, it's taken me a while to get this all straight in my head 15:31.000 --> 15:36.560 and make sure that we can move forward confidently. 15:36.560 --> 15:47.520 I've been trying very hard to remain true to my original idea, which was that this biology 15:47.520 --> 15:50.560 can't be impossible to learn. 15:50.560 --> 15:58.080 And any of the biology that underlies this phenomenon is worth learning. 15:58.080 --> 16:02.320 And so in the case of virology, I think all of us are just going to have to peel back 16:02.320 --> 16:11.040 our roll up our sleeves a little bit and learn some of this biology that they purport to 16:11.040 --> 16:16.600 have a monopoly on, that they purport to understand so well that you're just going to have to 16:16.600 --> 16:23.280 take their word for it, that they can't find any of this stuff in 2019 and before, 16:23.280 --> 16:26.280 but they sure can find a lot of it now. 16:26.280 --> 16:30.840 And that means that it's something going around and around and around and it couldn't 16:30.840 --> 16:34.060 be anything else. 16:34.060 --> 16:37.440 There are a few things in the background that we have to take and keep in mind. 16:37.440 --> 16:40.000 And there are a few things that we haven't addressed at all. 16:40.000 --> 16:45.680 And so it's just a real, it's a real mess right now. 16:45.680 --> 16:51.200 And so what they would like more than anything else for us to do is to rush as fast as we 16:51.200 --> 16:56.040 can thinking we know where we're going and thinking we know where the opening is to the 16:56.040 --> 16:57.040 cave. 16:57.040 --> 17:01.320 And we've got to keep remembering that we've got to keep our flashlights forward and we 17:01.320 --> 17:02.880 just got to keep working. 17:02.880 --> 17:11.560 It can't be distracted by the things that are going on, even if it is that you get fired 17:11.560 --> 17:26.800 or you get laid off or you get unexplainably your position is discontinued. 17:26.800 --> 17:34.280 And you are offered a non-disclosure agreement in exchange for $8,000. 17:34.280 --> 17:42.760 And you're left to kind of decide did I really put my neck on the line and put on 17:42.760 --> 17:55.240 the t-shirt and say that I was fighting for the truth and then now all of this hard work 17:55.240 --> 18:04.480 and sacrifice was worth a non-disclosure agreement and $8,000. 18:04.480 --> 18:08.640 I decided not to do that. 18:08.640 --> 18:13.760 And now I'm guessing I'm at the mercy of the internet for a little while. 18:13.760 --> 18:20.760 And we're going to try and make this work because I think actually this biology could 18:20.760 --> 18:23.600 save our kids. 18:23.600 --> 18:35.280 I actually think that if we can work through the literature and find truth in it, especially 18:35.280 --> 18:44.800 pre-COVID truth, and we can find biologists that we're working toward what they thought 18:44.800 --> 18:52.600 was the truth, I think we can find something that will be solid enough to stand on, that 18:52.600 --> 18:58.560 we can hand down to our children as an understanding of what these people lied about, what these 18:58.560 --> 19:02.080 people misled us about. 19:02.080 --> 19:08.000 And I think you're going to find that the illusion goes back farther than we thought. 19:08.000 --> 19:13.240 I think that's why so many of the interesting players in this are not like your typical 19:13.240 --> 19:21.400 players, like people who dropped out of grad school to start working on the Human Genome 19:21.400 --> 19:29.200 Project and then had like patents and companies and companies and ultra success. 19:29.200 --> 19:37.960 And then their boss went to the White House and was involved in the writing of $65 billion 19:37.960 --> 19:41.000 proposal for Warp Speed vaccines. 19:41.000 --> 19:49.920 And now he's been involved from the very beginning with objecting to the PCR test, but not PCR 19:49.920 --> 19:57.720 testing, and then all kinds of other random appearances for the RNA not being pure and 19:57.720 --> 20:06.920 then the RNA code on optimizing being responsible for different G quadriplexes and then came 20:06.920 --> 20:14.280 out later to be the double stranded DNA and the whole time this guy's been right there. 20:14.280 --> 20:19.920 Why has he been right there when his whole life is right now pot genetics and trying 20:19.920 --> 20:28.520 to, I don't know, I guess take over the commercial pot market by controlling genetics and virons 20:28.520 --> 20:37.640 and screening or providing testing materials for pot growers that will eventually become 20:37.640 --> 20:43.800 required by the government or some other scam. 20:43.800 --> 20:48.720 I'm sorry, but this is just this game has got to end. 20:48.720 --> 20:57.840 And these people, whether you like it or not, are not playing a small, tiny game. 20:57.840 --> 21:03.680 This is a game for all the marbles is for our grandchildren. 21:03.680 --> 21:08.720 That's why it seems so ridiculous. 21:08.720 --> 21:12.360 That's why it's being fought on Twitter. 21:12.360 --> 21:19.200 Do you think that Kevin McCurnan could have a real discussion about this and a group of 21:19.200 --> 21:23.960 people that all kind of knew about this stuff? 21:23.960 --> 21:28.960 Or would it would it really always have to be like him and Meryl Nast and then Andrew 21:28.960 --> 21:36.360 Kaufman and Christine Massey. 21:36.360 --> 21:42.800 The one time that we had a conversation a year ago, he couldn't really object to it. 21:42.800 --> 21:51.360 As you will see in tonight's recap, he just basically spit out exactly what I'm pointing 21:51.360 --> 21:55.640 out when everybody should be pointing out. 21:55.640 --> 22:04.240 That by making an infectious RNA and significant quantities, you could seed a coronavirus pandemic 22:04.240 --> 22:09.680 that was highly homogenous across many different places and maybe even would be highly stable 22:09.680 --> 22:11.440 for a little while. 22:11.440 --> 22:15.840 Yes, it's possible that clones transmit. 22:15.840 --> 22:20.760 I never said that they wouldn't. 22:20.760 --> 22:26.920 My main argument was that infectious clones and the whole principle of how they should 22:26.920 --> 22:33.520 or theoretically work is that you can create a much more homogenous viral stock that allows 22:33.520 --> 22:35.840 you to make a higher quantity of it. 22:35.840 --> 22:40.680 In fact, an infinite quantity of it is really only dependent on how to investment. 22:40.680 --> 22:45.760 If you wanted to do Jurassic Park, you could just do PCR construction of all the six or 22:45.760 --> 22:53.480 seven strands of DNA that you're going to use to ligate together and derive your RNA 22:53.480 --> 22:54.480 from. 22:54.480 --> 23:05.040 I mean, this guy's the limit if the idea is to just do it right. 23:05.040 --> 23:12.320 I'm just kind of, it's just kind of sad because this is where we're at, right? 23:12.320 --> 23:18.320 We have to protect our children from liars. 23:18.320 --> 23:24.800 Liars like Paul Offit, liars like Tony Fauci, liars like all of the people that work at 23:24.800 --> 23:31.200 the top levels of these pharmaceutical companies that know better. 23:31.200 --> 23:35.360 And we also have to protect our children from a bunch of people who don't know any better 23:35.360 --> 23:41.080 who aren't wise enough, smart enough, or I've turned the other way. 23:41.080 --> 23:44.280 In the military, they have caught what is called a lookaway doctrine. 23:44.280 --> 23:48.320 You know, if you're going to waterboard people, just let me close the door and get out of 23:48.320 --> 23:50.840 here before you start doing it. 23:50.840 --> 23:57.240 There are a lot of people involved in this little mystery that are also involved in that 23:57.240 --> 23:58.340 way. 23:58.340 --> 24:00.720 They know exactly how dangerous this was. 24:00.720 --> 24:04.980 They knew exactly how nasty this was going to get. 24:04.980 --> 24:08.800 And they knew that if they were going to get through this, they were going to break 24:08.800 --> 24:18.480 some eggs, but you know, make no mistake about it, ladies and gentlemen, breaking eggs 24:18.480 --> 24:26.960 is also going to require people on Twitter that have no business being on Twitter, talking 24:26.960 --> 24:32.400 about coronaviruses and fidelity of coronavirus replication and all this other stuff in their 24:32.400 --> 24:41.160 spare time while they're running companies on the genetics of weed. 24:41.160 --> 24:48.520 I mean, we are very, very, very close to exposing a group of people who some way or another 24:48.520 --> 24:53.840 were involved in the containing of the narrative at the same time they were involved in the 24:53.840 --> 25:01.680 perpetuating of the worst case scenario, making absolutely sure that the existence of the transmission 25:01.680 --> 25:07.400 of a novel virus was never questioned. 25:07.400 --> 25:15.200 And think about how strange it is that Eric Lander's former protege is the guy who was 25:15.200 --> 25:21.040 right there from the beginning, from the PCR objection to the RNA purity objection to the 25:21.040 --> 25:27.920 G quadriplex objections and the codon optimizing objections to the frame shifting objections 25:27.920 --> 25:32.920 all the way up to the double stranded DNA objections and what's really weird about the 25:32.920 --> 25:40.560 objections about the clones and about the proofreading in coronaviruses is that this 25:40.560 --> 25:46.360 thing that they recommended really, really early, that Cina Bavari predicted that they 25:46.360 --> 25:51.040 would use very, very early, that Rick Brighton knew that they were going to use very, very 25:51.040 --> 25:59.680 early, that Paul Cantrell knew that they were going to use very, very early, Remdesivir 25:59.680 --> 26:05.040 messes with the proofreading of coronavirus. 26:05.040 --> 26:08.240 So the proofreading of coronavirus is actually pretty significant. 26:08.240 --> 26:18.360 We need, we need to have high fidelity coronavirus replication that can be interfered with by 26:18.360 --> 26:25.840 Remdesivir and other antivirals so that the, the subtle balance between the mutagenesis 26:25.840 --> 26:32.400 in the quasi-species swarm and the stability of it is disrupted and you get this collapse 26:32.400 --> 26:35.240 of genetic stability. 26:35.240 --> 26:36.800 That's the story that they tell. 26:36.800 --> 26:38.720 That's the story that Mark Denison tells. 26:38.720 --> 26:41.800 That's the story that Cina Bavari told in his paper. 26:41.800 --> 26:44.200 That's the story that Ralph Barrick has told. 26:44.200 --> 26:50.760 And that's the story that is told when you argue that coronaviruses are very, very stable. 26:50.760 --> 26:56.000 You're arguing for Remdesivir's mechanism of action. 26:56.000 --> 26:57.000 Coincidence? 26:57.000 --> 26:59.000 Yeah, maybe. 26:59.000 --> 27:00.000 Believe what you want. 27:00.000 --> 27:01.000 What I want is high morbidity. 27:01.000 --> 27:02.000 I want people to complain. 27:02.000 --> 27:03.000 So what do I do? 27:03.000 --> 27:04.000 I go to Des Moines. 27:04.000 --> 27:05.000 Ladies and gentlemen, the people on the screen, I have nothing against Des Moines. 27:05.000 --> 27:06.000 I live there for four years. 27:06.000 --> 27:07.000 I go to Des Moines. 27:07.000 --> 27:08.000 I infect a couple of sentinel cases. 27:08.000 --> 27:09.000 I go to Seattle. 27:09.000 --> 27:12.000 I infect a couple of cases there. 27:12.000 --> 27:14.000 I go to North Carolina. 27:14.000 --> 27:15.000 I go to Wisconsin. 27:15.000 --> 27:23.000 What I'm doing is I'm using a dispersion methodology to be able to infect sentinel cases with a highly 27:23.000 --> 27:27.760 morbid condition. 27:27.760 --> 27:37.760 So I think what I want to do tonight is just quickly put up for a study hall. 27:37.760 --> 27:51.000 I've got this video queued up, a couple videos queued up on coronavirus replication, but 27:51.000 --> 27:56.640 I just want to look at... 27:56.640 --> 28:02.040 I just want to look at a couple things first to make sure that we still cover what we're 28:02.040 --> 28:03.040 really covering here. 28:03.040 --> 28:12.080 Let me escape out of this and start the new deck right away. 28:12.080 --> 28:19.200 I probably should just go really fast through it to bring everybody up to speed, but I hope 28:19.200 --> 28:20.800 you don't mind if I do that. 28:20.800 --> 28:28.640 So first of all, make sure you remember that one of the big, big, big things that is obvious 28:28.640 --> 28:35.240 to me as we move forward and we watch all these people scream and yell about how bad 28:35.240 --> 28:40.680 Jonathan Couey is for putting people's names on there or how bad it is that Jonathan Couey 28:40.680 --> 28:45.400 doesn't remember why somebody blocked him or whatever it is that they're crying about 28:45.400 --> 28:46.400 now. 28:46.400 --> 28:51.040 The one thing that you will remember and what I have been calling for more than a year now 28:51.040 --> 28:55.800 is that all these people will ignore this slide. 28:55.800 --> 28:59.960 You could be wrong about the clones, but right about this, and it wouldn't really matter 28:59.960 --> 29:01.360 now would it? 29:01.360 --> 29:03.840 And I assure you that I'm right about this. 29:03.840 --> 29:07.880 Intramuscular injection of any combination of substances with the intent of augmenting 29:07.880 --> 29:13.440 the immune system is dumb and transfection in healthy humans is criminally negligent. 29:13.440 --> 29:18.920 And by having me and you and all these people on Twitter that are trying to defend me, 29:18.920 --> 29:25.120 thank you very much, arguing about clones and whether or not the viral swarm is significantly 29:25.120 --> 29:30.200 diverse enough to support the idea that clones are different than the regular virus. 29:30.200 --> 29:34.440 This is all nonsense. 29:34.440 --> 29:38.400 This is all nonsense because this is true. 29:38.400 --> 29:43.560 This is all nonsense because this is true. 29:43.560 --> 29:49.120 And as long as they keep us occupied about whether or not J is right or wrong or whether 29:49.120 --> 29:54.720 it's okay to believe in J, then we're not going to we're not going to talk about this 29:54.720 --> 29:56.280 stuff. 29:56.280 --> 29:59.400 This is my most important message. 29:59.400 --> 30:04.880 This is the only message that I want to get to our children because if our children understand 30:04.880 --> 30:11.640 the big picture biology that underpins this statement, who cares about clones? 30:11.640 --> 30:17.040 Who cares how they do RNA virology and who cares if somebody with a Poch genetics company 30:17.040 --> 30:22.200 really does understand how they do coronavirus biology or not? 30:22.200 --> 30:31.680 Clearly he's being encouraged not to understand it because he is neglecting the very prominence 30:31.680 --> 30:40.440 of infectious clones from the early 90s on and the almost universal praise of how the 30:40.440 --> 30:45.640 development of infectious clones and their production has allowed the expansion of the 30:45.640 --> 30:51.120 understanding of how viruses work. 30:51.120 --> 30:56.400 We would have to be fools to believe that infectious clones weren't involved in this. 30:56.400 --> 31:02.400 They are the bedrock foundation methodology of all RNA virology, especially RNA virology 31:02.400 --> 31:08.520 around viruses that are difficult or impossible to culture for whom no infectious material 31:08.520 --> 31:12.800 is available. 31:12.800 --> 31:20.000 And so don't be fooled by somebody who can put up a few papers and then say a few sentences 31:20.000 --> 31:23.440 about why that paper makes them wrong. 31:23.440 --> 31:27.600 The time I've ever followed up on one of these papers, it doesn't really demonstrate 31:27.600 --> 31:36.520 what he says it demonstrates except for in a very sliver kind of way, but it's not addressing 31:36.520 --> 31:41.520 the very big pictures of what the natural phenomenon, what does the natural phenomenon 31:41.520 --> 31:45.520 look like is exactly the phenomenon that they don't want to talk about. 31:45.520 --> 31:50.400 They want you to believe and that's what he would like you to believe that the infectious 31:50.400 --> 31:59.360 clone replicates the natural phenomenon and that is a hundred percent wrong. 31:59.360 --> 32:04.920 And so they have fooled us into solving this Scooby-Doo and that's exactly what McCurnan 32:04.920 --> 32:06.440 is on right now. 32:06.440 --> 32:11.720 He wants you to believe that I'm supposed to have the exact explanation with all the details 32:11.720 --> 32:16.520 about exactly why the person came into my house, what they were doing there, what they intended 32:16.520 --> 32:20.280 to steal and why they were stealing it. 32:20.280 --> 32:24.840 And that's not the way a crime works. 32:24.840 --> 32:32.280 And our society, my family and yours are victims of a crime. 32:32.280 --> 32:36.120 We don't have any obligation as victims of a crime to know all the details and have a 32:36.120 --> 32:39.240 good explanation for everything. 32:39.240 --> 32:47.960 And we certainly aren't under any obligation to accept the explanation of the people who 32:47.960 --> 32:52.280 probably did it or are working with them. 32:52.280 --> 32:56.800 But we were fooled into believing that a worst-case scenario could have happened and we were fooled 32:56.800 --> 33:01.440 into believing that we could solve them, that we were watching people solve a mystery. 33:01.440 --> 33:08.880 That Rand Paul and Tony Fauci were arguing because they were on two sides of this mystery. 33:08.880 --> 33:12.560 And with this Scooby-Doo, the principle of informed consent has been ignored for the 33:12.560 --> 33:13.800 duration of the pandemic. 33:13.800 --> 33:22.560 Here we are trapped on our couch watching the whole thing unfold on TV. 33:22.560 --> 33:28.960 And so we've talked about how the pandemic was created from years ago up until now with 33:28.960 --> 33:35.520 a team of people or peoples around the world and especially in Western society that were 33:35.600 --> 33:39.600 there specifically to coordinate and amplify the worst-case scenario. 33:39.600 --> 33:47.440 The billions of people dead, the amyloid spike, the prion disease, the spike was killing cardiac 33:47.440 --> 33:50.840 cells, the whole nine yards. 33:50.840 --> 34:00.400 And the worst-case scenario was very cleverly tainted with aspects that they expected or 34:00.400 --> 34:04.120 feared would be a result of the transfection. 34:04.120 --> 34:07.280 They knew they were going to roll out the transfection so they knew that they could 34:07.280 --> 34:12.840 see this worst-case scenario with the worst-case scenarios of the transfection so that it would 34:12.840 --> 34:20.880 never really be clear to an unsuspecting and completely compliant populace that they were 34:20.880 --> 34:25.480 confounding the effects of the virus with the effects of the shot because you were given 34:25.480 --> 34:30.320 a viral protein and the viral protein turned out to be wicked. 34:30.320 --> 34:34.640 The people that were co-opted very early on were briefed that this was a national security 34:34.640 --> 34:40.360 thing and so you couldn't just talk about it. 34:40.360 --> 34:45.000 But if you go along with our little role here then maybe you can sell a book about Ivermectin 34:45.000 --> 34:52.760 or you can sell a book about hydroxychloroquine or you can sell a book about the lab leak. 34:52.760 --> 34:57.520 And in the end the toxicity of the virus, spike protein has been confounded with the 34:57.560 --> 35:03.520 toxicity of the transfection along with, of course, since we saw through this, since 35:03.520 --> 35:09.160 we saw through this little ruse a few years ago, a couple years ago, and we stopped believing 35:09.160 --> 35:12.760 this ruse. 35:12.760 --> 35:16.040 One of the reasons why we stopped believing this ruse was because we realized that the 35:16.040 --> 35:21.720 spike protein that's produced by a codon-optimized pseudo-uridine chemically altered RNA is not 35:21.720 --> 35:29.240 going to be anywhere structurally related in epitopes to that of one by the viral sequence. 35:29.240 --> 35:34.520 And we came to that conclusion after having Kevin McCurtain on our stream for two times. 35:34.520 --> 35:38.440 And so if the protein in three-dimensional structure is not equivalent to the protein 35:38.440 --> 35:43.040 that you're trying to build immunity to, chances are very good that the most damage 35:43.040 --> 35:48.040 associated immune epitopes there on that are also different. 35:48.040 --> 35:56.480 And so here we are, we know what the game was, we know how they did it. 35:56.480 --> 36:00.360 We know how they did it, we know they drove these mass casualty events and coordinated 36:00.360 --> 36:04.600 them in a way that made sure that nobody could question what actually happened. 36:04.600 --> 36:09.200 They co-opted a pre-selective group of narrative controllers and they probably brought more 36:09.200 --> 36:15.320 people on as the narrative went out of control as the compliance wasn't 100% with the uptake 36:15.400 --> 36:21.440 of the transfection, but a combination of background signal and nonspecific tests. 36:21.440 --> 36:24.640 And we don't have to figure that out for ourselves, we don't have to take everybody's 36:24.640 --> 36:27.160 word for it. 36:27.160 --> 36:32.720 And just because one person published a paper doesn't mean that that is the definitive 36:32.720 --> 36:39.720 Supreme Court evidence that now a scientific fact has been established, which is often 36:39.720 --> 36:43.520 how a lot of these rebuttal papers are presented. 36:43.960 --> 36:48.840 If you say somebody doesn't understand that the RNA, dependent RNA polymerase has a different 36:48.840 --> 36:55.320 replication capacity and then they cite a paper that doesn't immediately mean that 36:55.320 --> 37:00.920 that's the end of the discussion and that's really how this has gone from the very beginning. 37:00.920 --> 37:05.400 I think that they really thought that I was underprepared that I stumbled onto this idea 37:05.400 --> 37:08.960 and that's why the first three or four times that we had interaction with it, it was really 37:08.960 --> 37:11.080 pretty weak. 37:11.080 --> 37:16.280 And the one time we had an interaction live with my Kevin McCurnan and myself, I just 37:16.280 --> 37:21.440 kept kicking the ball back to him and at some point I just had to let him go because he 37:21.440 --> 37:27.040 just kept saying things that made it better. 37:27.040 --> 37:31.680 And so I think after that talk is when everything really peeled out of control and he decided 37:31.680 --> 37:36.520 that he had to make it appear as though he was slapping me down and used that stupid 37:36.520 --> 37:42.480 complicated strategy of his, you know, where he says, April back like six times and it 37:42.480 --> 37:47.360 forces everybody to Google it and so they all feel like fools and they don't want to admit 37:47.360 --> 37:52.920 that they don't understand what he's writing and then by not admitting that he's not teaching 37:52.920 --> 37:59.920 them anything, they let him get away with it, make it look like it's a consistent and consensus 37:59.920 --> 38:02.400 win on his part. 38:02.400 --> 38:08.880 And he refuses to acknowledge the fact that all of this stuff happened too. 38:08.880 --> 38:12.840 So there's actually really good reason to believe that there might not have been a real 38:12.840 --> 38:14.000 active spread. 38:14.000 --> 38:19.860 There's really good reason to be skeptical and that's part of what I find very funny 38:19.860 --> 38:24.720 about the one sub stack that actually dresses JJ's hypothesis in that sub stack. 38:24.720 --> 38:31.280 I think three times Kevin says maybe only twice that again, I'm not trying to decide 38:31.280 --> 38:36.040 here how much of the all-cause mortality should be attributed to COVID and how much 38:36.040 --> 38:42.120 of it should be attributed to bad ideas and man-demic. 38:42.120 --> 38:48.160 He makes the joke man-demic in his sub stack addressed to me from earlier this year, or 38:48.160 --> 38:55.720 maybe it's late last year, sorry, yes, it could be late 22 or early 23. 38:55.720 --> 39:02.800 And so I don't like to be accusatory, but in my perspective, we're dealing with someone 39:02.800 --> 39:05.080 who's behaving dishonestly. 39:05.080 --> 39:11.400 And from the very beginning, sort of a limited hangout kind of thing, giving as much, only 39:11.400 --> 39:16.560 as much information as to feel like he's giving a little bit, but never really bringing us 39:16.560 --> 39:17.840 all the way to the finish line. 39:17.840 --> 39:22.480 Are you telling me that Kevin McCurnan couldn't have imagined that there would be double-stranded 39:22.480 --> 39:26.800 contamination from the process two that he didn't know that they were going to use 39:26.800 --> 39:33.000 process to, that it was an option given the fact that Enovio was on BBC News with process 39:33.000 --> 39:36.600 two right behind them? 39:36.600 --> 39:38.480 I mean, how stupid do they think we are? 39:38.480 --> 39:42.160 How stupid does Robert Malone think we are that he doesn't know that process one and 39:42.160 --> 39:49.160 process two are very common differences that we knew that they made the joke on Twiv? 39:49.160 --> 39:55.240 Vincent Rancin-Yello talked about the amazing upgrade from nanograms to kilograms. 39:55.240 --> 39:59.880 I mean, come on, guys. 39:59.880 --> 40:05.840 And so these things all happened and they all want you to ignore the fact that they happened 40:05.840 --> 40:10.720 and they want you to continue to focus on a novel virus that's highly, high fidelity 40:10.720 --> 40:15.960 and that it can go around the world and change flavors repeatedly and with patterns. 40:15.960 --> 40:20.560 They want you to believe that passage of viruses gets you places. 40:20.560 --> 40:25.120 They want you to believe that when you stiff stuff together it gets you places. 40:25.120 --> 40:29.800 The worst-case scenario is that they made a lot of something and then they spread it 40:29.800 --> 40:30.800 around. 40:30.800 --> 40:33.880 That's the worst-case scenario. 40:33.880 --> 40:38.800 The worst-case scenario is that they made a lot of something that was pure and also toxic 40:38.800 --> 40:40.800 and fairly good at replicating. 40:40.800 --> 40:43.560 That would be awful. 40:43.560 --> 40:51.240 But it wouldn't be natural and none of the attributes of its spread would be natural 40:51.240 --> 40:55.000 because it would have started from a state that wasn't natural, a purity that wasn't 40:55.000 --> 41:01.440 natural and I can't understand why they insist on confounding this, ignoring this really 41:01.440 --> 41:03.640 easy thing. 41:03.640 --> 41:05.560 People asking me to write papers and stuff. 41:05.560 --> 41:10.160 It's like trying to write a paper about why cars use wheels. 41:10.160 --> 41:12.840 What does that mean? 41:12.840 --> 41:18.920 What does that mean when Alexandros Maranos and Kevin McCurnan say write up a paper? 41:18.920 --> 41:21.080 What does that mean? 41:21.080 --> 41:26.480 I'm telling you and it's a biological fact that the DNA can be copied with much higher 41:26.480 --> 41:33.280 fidelity than RNA and Kevin McCurnan even admitted it that you can make a lot of DNA 41:33.280 --> 41:36.720 and then you can make a lot of RNA from it and if you did that you would have a lot of 41:36.720 --> 41:37.720 genomic RNA. 41:37.720 --> 41:42.360 It'd be a little variable but it'd be a lot higher purity than anything you could ever 41:42.360 --> 41:49.400 achieve with any other method that we know and if you invested sufficient into the fidelity 41:49.400 --> 41:55.240 of it, use PCR methodologies, the state of the art technology, you could get purity levels 41:55.240 --> 41:58.240 heretofore on seed on earth. 41:58.240 --> 42:03.120 Distribute those everywhere and the signal would be brightest day. 42:03.120 --> 42:07.160 Not to mention the fact that you would have this all this leftover DNA that would also 42:07.160 --> 42:14.200 provide PCR signals because PCR is only looking for a very tiny amplicon so you'd 42:14.200 --> 42:20.920 have made tons of that amplicons too that you could distribute in sewers and in water 42:20.920 --> 42:25.280 and DNA is much more stable. 42:25.280 --> 42:30.840 That signal could potentially last for months and months depending on where you put it. 42:30.840 --> 42:35.640 Cruise ships could be positive for months if you had DNA that you distributed around. 42:35.640 --> 42:41.000 I mean come on ladies and gentlemen this is this is all obviously a joke thanks to 42:41.000 --> 42:44.120 James Giordano. 42:44.120 --> 42:49.080 Thanks to Kevin McCurnan, thanks to Kevin McCairn, thanks to Paul Catrell, thanks to 42:49.080 --> 42:56.920 Rick Bright, thanks to Tony Fauci, thanks to Ralph Barrick. 42:56.920 --> 43:01.060 You know one of the things that I think is really funny about Ralph Barrick is that actually 43:01.060 --> 43:07.700 I think he was a good guy and I think he's being co-opted. 43:07.700 --> 43:16.300 I want you to listen to this presentation that I think we are being set up to watch. 43:16.300 --> 43:18.460 That's all I'm going to say. 43:18.460 --> 43:24.300 I'm not sure if it's an authentic presentation, there are a couple times where it goes silent 43:24.300 --> 43:30.740 for like five or six seconds and I'm not sure it's an audio glitch, it could be an erasure 43:30.740 --> 43:35.020 and a couple times when it goes quiet they are talking about very key aspects of the 43:35.020 --> 43:37.660 infectious cycle. 43:37.660 --> 43:44.860 So this YouTube video was actually posted in a sub stack of Peter McCullough and this 43:44.860 --> 43:50.020 John O'leke or something like that and so that's how I found it and so I'm watching 43:50.020 --> 43:54.740 it for that reason but what I want you to remember is that this is in between SARS-1 43:55.740 --> 44:02.780 and Ralph Barrick may in theory, I want you to entertain this possibility, Ralph Barrick 44:02.780 --> 44:13.220 may still be an innocent bystander, a guy who's just part of the of coronavirus biology 44:13.220 --> 44:21.900 and he could still be in earnest thinking that coronaviruses could be one, a biotechnology 44:21.900 --> 44:32.780 to a pan vaccine technology and three, some kind of, I already said that, but some kind 44:32.780 --> 44:44.900 of future genetic modification technology so as they stand, some useful, then also modified 44:44.900 --> 44:50.980 and then also modified again I mean they were a huge potential in his mind and so I really 44:50.980 --> 44:56.380 think it's cool to listen to a video like this, even if it's altered, even if we're 44:56.380 --> 45:04.620 being tricked, I'm willing to bet that you're gonna hear enough of a guy who is fascinated 45:04.620 --> 45:12.500 by what he believes he is doing and I think he is doing it and that is studying synthetic 45:12.500 --> 45:14.180 viruses. 45:14.180 --> 45:19.020 So viruses that they can't culture, that they couldn't really get to grow in a lab and 45:19.020 --> 45:24.580 sustain themselves but has found another way to do it when we have these sequences 45:24.580 --> 45:29.780 and we decide on a consensus how can we do it and to really hear somebody explaining 45:29.780 --> 45:36.140 how they are developing this technology to assemble full coronavirus genomes, I'm confident 45:36.140 --> 45:43.260 it's gonna help us in terms of moving forward and really breaking the veil of this whole 45:43.260 --> 45:44.260 thing. 45:44.260 --> 45:53.380 Well, Carolina, you're gonna tell us about the synthetic genomics of SARS. 45:53.380 --> 45:59.820 So I also would like to thank the organizers for inviting me to talk, it's been wonderful. 45:59.820 --> 46:02.180 Now you are gonna be disappointed with the video here. 46:02.180 --> 46:05.380 Capture some new ideas that can be tried out on viruses. 46:05.380 --> 46:06.620 Well, this is not very loud. 46:07.620 --> 46:13.420 I'm gonna talk about today our basic short introduction into SARS biology. 46:13.420 --> 46:17.140 Talk about the synthetic resurrection and reconstruction of a variety of zoonotic SARS 46:17.140 --> 46:18.780 viruses and their applications. 46:18.780 --> 46:27.140 Okay, so already we're talking about synthetic reconstruction and he used the word resurrection. 46:27.140 --> 46:35.220 So resurrection would be from a sequence, from a partial sequence and it's very important 46:35.220 --> 46:40.740 to hear that because this is part of the reason why, again, clones are not just nothing. 46:40.740 --> 46:46.820 They're very special because the vast majority of coronaviruses cannot be easily cultured 46:46.820 --> 46:53.900 and when they are cultured, they only really barely culture it, they call it extremely 46:53.900 --> 46:59.660 low titer, but it's not really culturing that you can't make enough to share and it's 46:59.660 --> 47:00.660 not culturing. 47:00.660 --> 47:01.780 So they have a big problem. 47:01.780 --> 47:08.340 It's not just that they don't have, they don't have them in the right cell culture 47:08.340 --> 47:12.980 or it's not the right receptor, it's more complicated than that. 47:12.980 --> 47:19.580 And I think it comes down to the fact that infectious particles are oftentimes replication 47:19.580 --> 47:25.820 and competent and if you get at the heart of this, you're gonna find that that's really 47:25.820 --> 47:33.300 where this all peters out, where the rubber meets the road is right there and that's 47:33.300 --> 47:35.620 where they just do the hand wavy. 47:35.620 --> 47:38.620 And that's why the no virus people had so much to stand on. 47:38.620 --> 47:46.460 That's why I think they were so dangerous and I think that's why they were so confusing. 47:46.460 --> 47:52.020 But are we, do you feel the pendulum swinging back now, do you feel it? 47:52.020 --> 47:56.740 Where we were almost all the way to no virus and really exploring the possibility that 47:56.740 --> 48:00.820 wow, did they really only put clones in like four people? 48:00.820 --> 48:05.300 Like Sohomish County man and a couple other people and sequence those and then the rest 48:05.300 --> 48:09.380 is all nonsense, it's possible. 48:09.380 --> 48:16.420 But there's a whole possibility space that expands from that very tiny number of people 48:16.420 --> 48:23.540 that were infected with clones that never went anywhere to another medium possibility, 48:23.540 --> 48:29.140 which is that they actually did put a significant a number of and concentration of clones in 48:29.140 --> 48:34.340 several places so that several people or many hundreds of people would be infected with 48:34.340 --> 48:37.940 a very pure infectious RNA. 48:37.940 --> 48:42.660 And I think over the next few days, as we explore these papers together, you're going 48:42.660 --> 48:49.940 to find a preponderance of evidence as I have, which indicate that these RNAs all by themselves 48:49.940 --> 48:53.220 do some pretty cool stuff. 48:53.220 --> 49:05.460 And as far as most virology papers are concerned, these are better than or more reliable than viruses. 49:05.460 --> 49:09.860 And what they do and how they do it is more reliable and you're going to figure out that 49:09.860 --> 49:18.500 that's because of the purity of the RNA, the purity of genomic RNA that gets into each cell 49:18.500 --> 49:27.540 is several orders of magnitude higher than the amount of genomic DNA that gets into a cell 49:28.180 --> 49:30.740 during passage. 49:30.740 --> 49:35.780 And that's what's really extraordinary here. That's really what's extraordinary here. 49:36.420 --> 49:40.420 And therapeutic and vaccine design. 49:40.420 --> 49:45.220 I'll also touch on codons, the optimization as a way to attenuate SARS pathogenesis, 49:45.220 --> 49:49.940 but I certainly haven't gone to the depth of studies that Eckerd's group has done. 49:49.940 --> 49:55.620 And then I'll talk about rewiring SARS-Coronavirus transcription circuits as a way universal strategy 49:55.620 --> 49:59.380 to attenuate viral pathogenesis. 49:59.380 --> 50:03.060 So this is a schematic diagram of the SARS-Coronavirus particle. 50:04.020 --> 50:06.420 It's about 100 nanometers from diameter. 50:06.420 --> 50:09.940 It contains a helical nucleic acid inside. 50:09.940 --> 50:12.820 It contains a single-stranded plus polarity RNA genome. 50:14.020 --> 50:17.140 This nucleic acid structure is surrounded by a lipid bilayer. 50:17.780 --> 50:20.500 It contains several viral glycoprotein spikes. 50:20.500 --> 50:25.060 The important ones for today's talk include the M glycoprotein and the E protein, 50:25.060 --> 50:26.260 which are shown right here. 50:26.820 --> 50:30.900 These proteins are absolutely essential for maturation and release 50:30.900 --> 50:32.580 and the production of virus particles. 50:33.220 --> 50:34.260 So keep that in mind. 50:35.220 --> 50:39.540 The second viral protein of interest for today's talk is the escolycoprotein gene 50:39.540 --> 50:43.460 that's shown right here, which gives the virus its unique occurrence in the electron microscope. 50:44.740 --> 50:48.820 It's the viral passion protein that binds to the receptor to mediate, 50:48.820 --> 50:50.180 docking, and entry into the cell. 50:51.140 --> 50:55.460 It regulates tissue tropism, species specificity. 50:55.460 --> 50:58.340 It contains a large number of important neutralizing epitopes, 50:58.340 --> 51:00.900 and it's the principal component of protective immunity. 51:01.860 --> 51:08.340 Now, if you tear this virus particle apart and look at the viral genomic positive-stranded RNA, 51:08.340 --> 51:11.060 it's about 30,000 base pairs in length. 51:11.060 --> 51:14.180 It's the largest plus polarity RNA genomes in nature. 51:14.180 --> 51:17.940 The first 20,000 base pairs are shown to encode the replicase proteins. 51:17.940 --> 51:22.340 So the job it is to replicate the genome, replicate the genome, 51:22.340 --> 51:26.740 and express sub-genomic messenger RNA that encode these downstream open reading frames. 51:31.540 --> 51:33.540 Here's the first blank space. 51:33.540 --> 51:37.460 Now, these downstream morphs encode the structural genes like the SEM and 51:37.460 --> 51:40.980 nucleocapsid protein that I just mentioned that were present in the virion 51:40.980 --> 51:43.220 in a variety of accessory morphs. 51:43.220 --> 51:48.020 To get these expressed in cells, you find a nested set of sub-genomic messenger RNA. 51:48.580 --> 51:52.580 They're ranged from the three prime end of the genome so that all the sequences in the smallest 51:52.580 --> 51:56.260 message are in the next largest message and so on and so forth. 51:56.260 --> 52:01.620 This organization allows different open reading frames to be placed at the five prime ends of 52:01.620 --> 52:06.100 different messenger RNAs so they can be efficiently translated and expressed in cells. 52:06.820 --> 52:07.700 Now, the trans... 52:09.460 --> 52:11.140 Here, again, is another dropout. 52:13.540 --> 52:13.940 I don't know. 52:13.940 --> 52:16.740 How 70 nucleotides shown here are these little blue boxes. 52:18.260 --> 52:21.460 That is derived from the five prime end of the genome. 52:21.460 --> 52:25.620 So these are discontinuous stretches of RNA that are joined going transcription. 52:25.700 --> 52:29.940 And these little red box elements right here are absolutely essential 52:29.940 --> 52:32.420 for mediating the joining of these two sequences. 52:33.380 --> 52:36.020 This will become very important later on in the talk. 52:36.020 --> 52:36.980 So keep the... 52:36.980 --> 52:41.940 Okay, so what he's explaining here just to kind of to bring you up to speak. 52:41.940 --> 52:44.020 You can't really see it here and I apologize. 52:44.020 --> 52:46.740 I told you it was going to be a bad... the graphics are bad. 52:48.660 --> 52:51.060 It's weird. This was uploaded last year. 52:51.060 --> 52:52.740 I can't find another copy of it. 52:53.620 --> 52:58.820 There's those two things that were edited out there or blanked out in the beginning there. 52:58.820 --> 53:02.100 I can't tell but it sure feels like there was an erasure there. 53:02.660 --> 53:07.700 And he's talking about these TRSs which are translation regulatory sequences. 53:08.660 --> 53:12.660 They are small sequences that are repeated throughout the genome. 53:13.380 --> 53:17.140 And it's not very clear to me upon reading the genome... 53:17.780 --> 53:24.660 or sorry, the literature exactly how these transcriptional regulatory elements 53:25.380 --> 53:26.660 interact with one another. 53:26.660 --> 53:36.900 But it feels like, if I can give you my best impression, it feels like that these TRS segments 53:37.620 --> 53:44.740 are repeated because the RNA molecule has a three-dimensional interaction with itself 53:45.300 --> 53:50.260 and makes it not so that this part gets looped over to this part. 53:50.820 --> 53:56.580 And so then the RNA-dependent RNA polymerase can make a very short RNA that goes bloke 53:56.580 --> 53:57.860 and then it goes right to here. 53:58.740 --> 54:01.540 And it goes bloke and then it goes right to here. 54:01.540 --> 54:06.020 And so every time it gets to a translational regulatory element, 54:06.660 --> 54:12.420 there's the potential for this part of the five prime end to be looped in over here. 54:12.420 --> 54:16.020 And so then the RNA-dependent RNA polymerase that's translating it can 54:16.980 --> 54:19.300 to make this sub-genomic RNA. 54:19.300 --> 54:21.620 And that's probably completely wrong. 54:21.620 --> 54:26.180 But I do know that the translation regulatory elements are all the same. 54:27.140 --> 54:33.940 And it is from these spots in the genome that the recombination is most likely to occur. 54:35.780 --> 54:40.020 And one of the ways, and Ralph Berwick will explain it in this talk later, 54:40.100 --> 54:44.420 that they thought to make an attenuated virus that could not 54:45.220 --> 54:49.860 de-attenuate by recombining with other viruses in the wild 54:50.660 --> 54:56.980 was to artificially change the translation regulatory segments of the artificial genome 54:57.940 --> 55:04.980 so that this virus, when co-infecting a cell, would have translation regulatory elements that 55:04.980 --> 55:09.700 were incompatible with the genomes that would be present in the wild. 55:09.700 --> 55:17.620 And therefore they could not rearrange their genome to acquire genes from wild coronaviruses. 55:17.620 --> 55:23.460 So what he's explaining here is an aspect of the coronavirus genome that if you take their 55:23.460 --> 55:29.700 word for it, as I'm doing right now, is part of the regulatory mechanism by which the 55:30.660 --> 55:36.500 entire signal genome actually gets translated into these sub-genomic RNAs that we've been talking 55:36.500 --> 55:41.540 a lot about and that we understand are in abundance of several orders of magnitude more 55:41.540 --> 55:47.300 abundance than the full genome, whenever they try to find the total RNA in any of these preparations. 55:47.300 --> 55:53.940 I hope that helped a little bit, especially because you can't see Jack in this sort of blurry thing. 55:53.940 --> 55:58.180 But we are listening to the great RB, so it's pretty fun. 55:58.740 --> 56:04.020 The concept of TRS elements, a little red box element, is being essential for regulation of 56:04.020 --> 56:12.420 messenger RNA synthesis. Now, let's talk about synthetic genomics in the context of a platform 56:12.420 --> 56:17.780 to control emerging infectious diseases. If you think about emerging viruses, they have tremendous 56:17.780 --> 56:23.940 potential to cause high morbidity and mortality. They have tremendous potential to cause high 56:24.020 --> 56:31.220 morbidity and high mortality, meaning that there are small examples of this happening. But again, 56:31.220 --> 56:38.100 it's the pandemic potential is different. I'm sure that if you had enough clone of a clone 56:38.100 --> 56:42.660 of an RNA like this and you put it in somebody's lungs, yeah, or squirted it in some, 56:42.660 --> 56:49.940 an animal's nasal passages, you'd have some problems. If you put that RNA in a cell culture and then 56:49.940 --> 56:54.580 passage it a few times, you're going to have a lot of RNA in every cell. It's all going to be 56:54.580 --> 56:58.820 the same. It's all going to do its viral thing. It's all going to package a bunch of stuff up. 56:58.820 --> 57:05.780 You're going to have a huge concentration of highly homogenous RNA particles, as far as we can tell, 57:05.780 --> 57:12.500 viral particles, exosomes, whatever, that you would not have if you just started with a sample from 57:12.500 --> 57:17.300 the bat and then started passageing it and tried to make a lot. And that's the point. 57:21.220 --> 57:26.900 Let's let it keep playing here. Maybe I should just quick, just let me give you this one thing here. 57:26.900 --> 57:31.220 I had this queued up and then I just, I just realized we were going way, I was going way off 57:31.220 --> 57:43.060 balance here. But here's a mini review from a virology in 1994, infectious transcripts, 57:43.060 --> 57:50.020 CDNA clones of RNA viruses. Recombinant DNA technology makes it possible to analyze and modify 57:50.020 --> 57:55.540 genomes at the molecular level and thus gain deeper insight into their organization and expression. 57:55.700 --> 58:01.140 Oops, sorry. In this, in this respect, viruses because of the small size of their genome are 58:01.140 --> 58:06.820 particularly amenable to such investigations. In spite of this, the study of molecular biology of 58:06.820 --> 58:13.860 non-retroviral RNA viruses has long been hampered by the fact that these viruses do not encompass a 58:13.860 --> 58:20.740 DNA intermediate step in their replication cycle. Therefore, since to date, the extremely varied 58:20.740 --> 58:26.020 and powerful molecular biology techniques aimed at modifying nucleic acids have been directed 58:26.020 --> 58:31.780 essentially at DNA substrate, new molecular tools had to be developed. The possibility of 58:31.780 --> 58:42.260 obtaining infectious clones as CDNAs or as in vitro transcribed RNA copies, which is what an RNA 58:42.260 --> 58:50.900 infectious clone is. Corresponding to the genomes of RNA viruses has greatly enhanced the 58:50.900 --> 58:56.100 potential of investigations. Indeed, they can facilitate studies of viruses that are present 58:56.100 --> 59:07.940 only in low titers in infected cells or whose isolation is problematic. Interesting. That's from 59:08.340 --> 59:20.820 1994. Let's go to 2019 from Cina Bavari and Alison Totura, a former postdoc of Ralph Barrick. 59:22.340 --> 59:30.100 Section 2, 2.1, reverse genetic systems. Advances in the study of highly pathogenic coronaviruses and 59:30.100 --> 59:35.460 potential pan-coronavirus drug tech candidates partially depends on the technology to genetically 59:35.460 --> 59:40.180 manipulate coronaviruses to probe mechanisms of viral pathogenesis and antiviral drug activity. 59:40.820 --> 59:46.740 Reverse genetic systems synthetically generate viruses from known viral sequences. In situations 59:46.740 --> 59:52.500 who wear clinical isolates of infectious material unavailable and due to restrictions for collecting 59:52.500 --> 59:57.140 patient samples, shipping infectious materials, availability of containment laboratories, 59:57.140 --> 01:00:03.220 reverse genetic systems, provide the essential research materials for studies on viral pathogenesis 01:00:03.300 --> 01:00:09.860 in model development. Prior to the SARS pandemic, robust reverse genetic systems to manipulate the 01:00:09.860 --> 01:00:17.140 genomes of Scarsco V2s, sorry, of coronaviruses had already been developed by systematic assembly 01:00:17.140 --> 01:00:23.060 of cDNA consents to full-length infectious clones, allowing precise and genetic targeted 01:00:23.060 --> 01:00:29.220 manipulation of viral genes. More importantly, to me, infectious clones allow the creation of 01:00:29.220 --> 01:00:36.900 near homogenous viral stocks where the traditional viral stocks are prepared by 01:00:36.900 --> 01:00:44.660 amplification of infectious material in cell culture. Infectious materials, that's the stuff 01:00:44.660 --> 01:00:51.460 they take from bats, that's the stuff they take from sick people. Over many passages, strategies 01:00:51.460 --> 01:00:56.580 to build reverse genetic systems were rapidly applied to both SARS and MERS within the first 01:00:56.580 --> 01:01:03.700 year of identifying these viruses. In addition to reconstructing epidemic, 01:01:03.700 --> 01:01:09.780 epidemic strains of SARS-CoV-2 with reverse genetic systems allow targeting of mutations 01:01:09.780 --> 01:01:15.860 to specific viral genes and assembly of these virus when infectious material is not available. 01:01:15.860 --> 01:01:28.260 So they've seen pretty significant for RNA virology in general, but for some reason or another, a guy who 01:01:30.980 --> 01:01:35.220 was at the White House when he was what, 25 or something like that? I don't know, 01:01:35.220 --> 01:01:40.420 how old was he when he was at the White House for the first time and probably because of his dad, 01:01:40.500 --> 01:01:49.300 maybe, I don't know, but and then his, he drops out of grad school, goes to work for Eric Lander, 01:01:50.260 --> 01:02:01.300 Eric Lander's in the Biden White House, running stuff for the mRNA, and this guy's involved from 01:02:01.300 --> 01:02:06.660 the very beginning from the PCR shot all the way through all the objections that were ever made 01:02:06.660 --> 01:02:17.220 about the transfection and titrating them one by one and we're supposed to believe that 01:02:17.220 --> 01:02:21.860 clones are just like everything else. It's no big deal. Clones don't do anything. Clones are 01:02:21.860 --> 01:02:31.780 chemtrails stupid. It's pretty phenomenal, really, if you think about it. And severe economic 01:02:31.780 --> 01:02:37.780 hardship, good examples are HIV, the 1918 flu, far as coronavirus, H5N1, 01:02:37.780 --> 01:02:43.220 chicken and guinea virus, probably most recently. Now it's clear that zoonotic introductions are 01:02:43.220 --> 01:02:50.260 increasing and a large number of these viruses have large reservoir pools of animal strains 01:02:50.260 --> 01:02:56.420 like SARS and H5 that are maintained as sort of heterogeneous swarms of pools of viruses 01:02:56.420 --> 01:03:01.220 of which someone may actually emerge in the future to cause epidemic disease in humans. 01:03:01.860 --> 01:03:07.140 This raises a conundrum in terms of how do you protect the public health against the future 01:03:07.140 --> 01:03:14.340 occurrence that is hard to predict. I wonder how we would differentiate between a pathogen in bats 01:03:14.340 --> 01:03:21.460 that doesn't make bats sick. How would we differentiate that from a immune signal? 01:03:22.900 --> 01:03:31.380 Like let's say that the bats physiology is to emit coronaviruses. Let's say that a lot of 01:03:31.860 --> 01:03:44.180 mammals emit self replicating RNA packaged in a lipoprotein coat. Just just imagine that for a 01:03:44.180 --> 01:03:51.300 second. Is that crazy? Is that a crazy idea? Are we crazy? Are we a little crazy to assume 01:03:52.100 --> 01:03:57.780 that these RNA molecules coated in a lipid dental, a lipid protein, a lipoprotein coat, 01:03:58.020 --> 01:04:03.060 aren't we a little? Isn't it a little weird to assume that these guys are just all independent 01:04:03.060 --> 01:04:12.740 actors that are out of the control of the bats, out of the influence of the bats, 01:04:12.740 --> 01:04:20.660 out of the hands of the, despite the bats best effort, these coronaviruses are thriving inside 01:04:20.660 --> 01:04:23.620 of the bat population. Why does it have to be like that? 01:04:28.260 --> 01:04:31.300 Is it, is it really like that? Are we sure it's like that? 01:04:34.340 --> 01:04:41.460 Are we sure that Simeon virus 40 was really a virus that the monkeys body really wanted to get 01:04:41.460 --> 01:04:47.300 rid of but couldn't get rid of? Are we assuming that the pangolans have coronaviruses that they 01:04:47.300 --> 01:04:54.020 don't want that they're not supposed to have but they're just there because despite their best 01:04:54.020 --> 01:05:05.060 efforts, that's a huge assumption, isn't it? As opposed to it being like a signal or an offgassing 01:05:05.060 --> 01:05:10.660 or a noise, I like a signal. I like the idea of it being a signal. 01:05:11.460 --> 01:05:15.940 I don't like the idea so much of it being a pathogen anymore. 01:05:17.940 --> 01:05:22.900 But I definitely like the idea that Ralph Barrick thinks it's a signal and thinks that what he's 01:05:22.900 --> 01:05:27.620 working on is important and what he's working on is something that's worth understanding and that 01:05:27.620 --> 01:05:33.540 he's made a major breakthrough by figuring out how to make these full genomes and start there. 01:05:34.340 --> 01:05:40.820 I think it's possible. I think your your sliver of understanding, your sliver of specialty could 01:05:40.820 --> 01:05:47.060 be so great that you could get so fired up about making machine guns that you just don't realize 01:05:47.060 --> 01:05:55.460 that, wow, I made really, I mean this machine gun is pretty amazing and I'm proud of all the the 01:05:56.340 --> 01:06:02.100 the ingenious ideas I put into it but I'm a little disappointed with what it did to them all. 01:06:03.060 --> 01:06:05.700 That's possible. It's totally possible. 01:06:08.420 --> 01:06:13.220 You know, I was a clock maker but then I got into guns and I just figured out this thing and it 01:06:13.220 --> 01:06:21.460 was super cool and and then somebody took it to a high school but you could still have your you 01:06:21.460 --> 01:06:27.060 know if you were really hyper focused on something if you really were interested in making something 01:06:27.060 --> 01:06:31.140 cool out of coronavirus is because somebody said that something cool could be made out of 01:06:31.140 --> 01:06:35.940 coronaviruses and you're a clever guy maybe that's who Robert over there that's who Ralph 01:06:35.940 --> 01:06:44.660 Berwick is. That's why he's trying to attenuate them. That's why he's trying to find combinations 01:06:44.660 --> 01:06:49.220 of monoclonal antibodies that will neutralize all of them. 01:06:52.580 --> 01:06:57.780 That's why he's trying to change their translation regulatory sequences so that they can't recombine 01:06:58.660 --> 01:07:07.060 with with natural viruses and that they can be permanently attenuated. These are all laudable 01:07:07.060 --> 01:07:14.180 pursuits if viruses are real and and if they are potentially a biotechnology that could be harnessed 01:07:14.180 --> 01:07:22.420 for good. A endogenous signal that could be that could be used and harnessed as a as a tool. 01:07:22.420 --> 01:07:31.380 Why not? Sounds great. I think Ralph Berwick might be innocent. I think we might be led to believe 01:07:31.380 --> 01:07:35.940 that in this little Scooby-Doo this is the guy we're supposed to unmask. This is the bad guy right 01:07:35.940 --> 01:07:42.900 here. Look at him. I don't think so. How do you develop drugs and therapeutic against an unknown 01:07:42.900 --> 01:07:47.940 from this heterogeneous swarm of variant strain that will actually work against the unknown that 01:07:48.020 --> 01:07:53.700 will emerge in the future? How do you target your scarce resources? We think platform technologies 01:07:53.700 --> 01:07:58.420 like synthetic biology, phylogenomics, structure modeling, high-throughput sequencing really will 01:07:58.420 --> 01:08:04.420 provide a platform and our goal is to use SARS coronavirus and its large pool of zoonotic viruses 01:08:04.420 --> 01:08:09.940 as a model to develop rapid response platforms to protect against this broader heterogeneous 01:08:09.940 --> 01:08:16.020 pool of variants. We also would like to develop strategies that would attenuate all family members 01:08:16.020 --> 01:08:20.580 so that you could rapidly develop vaccines for the public overall public health. 01:08:20.580 --> 01:08:25.300 And so that's funny because he's talking about rapidly developing vaccines. I wonder 01:08:25.300 --> 01:08:31.700 anywhere in this talk he's going to project a time frame. Is he going to say five years? 01:08:33.060 --> 01:08:37.140 Because if he says two years that would be very interesting, wouldn't it? If he says 01:08:37.860 --> 01:08:42.900 five years or he says one year anything that he says about development time will be interesting 01:08:42.980 --> 01:08:49.460 in 2007. This is going to be cool. Keep your ears open. Now a lot of beautiful work in Malek's 01:08:49.460 --> 01:08:55.940 lab as well as other labs here in China worked out the basic molecular epidemiology of the SARS 01:08:55.940 --> 01:09:01.060 coronavirus. The virus originated most likely in bat. It probably jumped into civets or humans 01:09:01.700 --> 01:09:07.780 and in that context set up a transmission cycle between civets and ratoon dogs and marketplaces 01:09:07.780 --> 01:09:13.140 and humans who frequented those marketplaces. Over time it's selected for strains that were 01:09:13.140 --> 01:09:17.540 more efficient at infecting human cells and recognizing the human ACE2 receptor for doctor 01:09:17.540 --> 01:09:24.420 human entry leading to the 2002-2003 epidemic which caused about 8,000 cases and 800 deaths. 01:09:26.500 --> 01:09:30.980 Now I want you to keep 8,000 cases and 800 deaths in mind as the 01:09:31.060 --> 01:09:44.740 as a certain quantity of clone. Let's call that one clone unit and so if you release one clone 01:09:44.740 --> 01:09:56.820 unit in a place in China and it can cover 8,000 trackable or suspected cases and 800 deaths 01:09:57.460 --> 01:10:08.260 with a few sequences. So let's just call that a single clone unit as 8,000 cases and 800 people 01:10:08.260 --> 01:10:16.820 dead so that we can think about what we would do if we had two or three clone units, if we had 01:10:16.820 --> 01:10:25.700 10 clone units released in four places. How many cases would that be? How far would it go? 01:10:26.900 --> 01:10:33.540 How much molecular change would occur per unit time in that scenario? I want you to start 01:10:33.540 --> 01:10:40.020 thinking about the possibility that that's what actually was done. The first time single clone 01:10:40.020 --> 01:10:48.340 unit this time maybe five clone units and the difference is that these five clone units were 01:10:48.340 --> 01:10:55.460 all so clones right they're the same so instead of having a SARS outbreak originate from a apartment 01:10:55.460 --> 01:11:04.820 building in China and ultimately end up somewhere in Toronto of all places which was probably part 01:11:04.820 --> 01:11:09.300 of an exercise again because they were elevated as people who knew what they were talking about 01:11:09.380 --> 01:11:18.420 at the beginning of this one and so with more clone units you could just have a bigger brighter 01:11:18.420 --> 01:11:26.820 signal that would be as homogenous as you wanted it to be and so interestingly enough when we were 01:11:26.820 --> 01:11:37.060 having this argument before Kevin McCurnan and I he brought up the idea that there should be some 01:11:37.060 --> 01:11:44.420 purity in the thing which I find curious and I just want to throw that up there for people who 01:11:44.420 --> 01:11:54.420 are listening and watching so this was from earlier this year something you know the things that he 01:11:54.420 --> 01:12:00.260 says that I ignore this is from a slide that I made for that I think it was in the in the last week 01:12:00.260 --> 01:12:06.180 of April the first week of May ask yourself why he's making six hour scooby-doo videos instead 01:12:06.180 --> 01:12:11.060 of downloading data from NCBI and trying to find a signal for his hypothesis in real data 01:12:11.060 --> 01:12:19.060 if the clones offer some advantage to Dr. Evil surely there is evidence of reduced mutation rates 01:12:19.060 --> 01:12:26.340 to prove this now interestingly I didn't want to make a big deal about this I went really fast 01:12:26.340 --> 01:12:30.820 through it because I'm a little scared it's going to disappear I'm a little scared that somebody's 01:12:30.820 --> 01:12:37.460 going to make an excuse for it to disappear but this is the paper from Alina Chan in May of 2020 01:12:37.940 --> 01:12:46.180 Alina Chan with the book Alina Chan that works at the Broad Institute that Eric Lander is the head 01:12:46.180 --> 01:12:54.420 of at MIT Eric Lander the former boss of Kevin McCurnan at the Human Genome Project ladies and 01:12:54.500 --> 01:13:01.860 gentlemen the one that worked at the White House for Biden yeah him now runs the Broad 01:13:01.860 --> 01:13:06.820 Institute or still runs the Broad Institute at MIT where Alina Chan works Alina Chan submitted 01:13:06.820 --> 01:13:14.820 this paper this pre-print to bio archive on May in 2020 and here she has nucleosides nucleotide 01:13:14.820 --> 01:13:22.820 substitutions in the genome of SARS-1 versus nucleotide substances substitutions in this genome 01:13:22.820 --> 01:13:29.860 of SARS-CoV-2 and lo and behold what is this it looks like there is evidence of reduced mutation 01:13:29.860 --> 01:13:34.660 rates at the beginning of the pandemic relative to the beginning of the pandemic of SARS 01:13:36.580 --> 01:13:45.060 hugely reduced mutation rates publicized and observed by none other than Alina Chan who was 01:13:45.060 --> 01:13:49.780 incidentally has never published this paper officially under peer review 01:13:50.500 --> 01:13:57.300 and instead accepted a position at the Broad Institute as a staff scientist and sold a book 01:13:57.300 --> 01:14:06.020 about the lab leak it's a pretty interestingly tiny circle of monkeys considering the fact 01:14:06.020 --> 01:14:17.300 that Kevin McCurnan just recently said that I gave him the George Webb treatment like I mean 01:14:17.300 --> 01:14:22.500 have we are really running out of characters here is it really that many people aren't backstage 01:14:22.500 --> 01:14:31.060 anymore or what like when we when we spike this football ladies and gentlemen it's going to be 01:14:31.060 --> 01:14:37.380 quite a spike okay it's going to be quite a spike let's go back to some more traditional virology 01:14:37.380 --> 01:14:43.140 with the Great Ralph Barrack if you do phylogenetic analysis of the isolates that were sequenced 01:14:45.620 --> 01:14:52.340 you find they fall into two broad categories these this line indicates the strains underneath 01:14:52.340 --> 01:14:58.020 that line were identified during the epidemic and you have early phase isolates primarily of 01:14:58.020 --> 01:15:04.260 January 2003 after a variety of super spreading events you had middle phase isolates shown here 01:15:04.260 --> 01:15:11.460 by this group of isolates and then following infected physicians who came to Hong Kong and 01:15:11.460 --> 01:15:16.100 resulted in a super spreading event that some transmitted the virus to the rest of the world 01:15:16.100 --> 01:15:22.100 these were considered late phase isolates now if anything is suited to a super spreader event 01:15:22.100 --> 01:15:27.780 it is somebody who is infected with an infectious clone because all their cells will have been 01:15:27.780 --> 01:15:34.020 infected with a genomic RNA that was identical which means when they start making this swarm 01:15:34.020 --> 01:15:42.180 of of slightly altered viral genomes and they make this huge collection of sub genomic RNAs 01:15:42.740 --> 01:15:47.860 the collection of proteins that they produce will be much more homogenous than anything 01:15:47.860 --> 01:15:53.860 that would have been generated from a typical cultured virus if you could culture it but you 01:15:53.860 --> 01:15:58.340 can't you see you see what's magical about this you see how beautiful this is 01:16:01.380 --> 01:16:06.340 and so Ralph knows that he's making something that's super cool Ralph knows that he's making 01:16:06.340 --> 01:16:14.180 something that is not natural but he's arguing that not only can we make something super pure 01:16:15.380 --> 01:16:22.580 but if we find a way to alter it so that it's not dangerous so that it's useful to us we can make 01:16:22.580 --> 01:16:31.060 huge quantities of that useful new synthetic virus from Ralph Barrack's perspective he is on 01:16:31.060 --> 01:16:38.420 the verge of tapping into a biotechnology that has almost limitless possibilities 01:16:41.060 --> 01:16:46.900 once he can figure out a way to attenuate them so that they replicate in a way that we want them 01:16:46.900 --> 01:16:53.140 to that they have no toxic parts to them that the most toxic parts of their replication process 01:16:53.140 --> 01:16:59.860 are minimized then potentially we would have the ultimate transfection tool the ultimate gene 01:16:59.860 --> 01:17:06.980 therapy tool the ultimate personalized medicine tool that's what he sees here that's what Ralph 01:17:06.980 --> 01:17:14.420 Barrack is chasing he's chasing a universal vaccine a universal transfection tool a universal gene 01:17:14.420 --> 01:17:22.580 therapy that's what he's teaching chasing here and he wants to use this diverse set of coronaviruses 01:17:22.580 --> 01:17:29.620 in the wild that's present and sustaining in the wild in these populations has a basis for understanding 01:17:29.620 --> 01:17:39.540 them if this biology is it checks out it's a very noble effort in my humble opinion it's not the 01:17:39.540 --> 01:17:48.820 Scooby-Doo bad guy we're looking for it's just not the vast pool of heterogeneous zoonotic strains 01:17:48.820 --> 01:17:53.540 however reside up here most of these were in fact none of these were ever actually successfully 01:17:53.540 --> 01:18:01.460 cultured they actually exist as sequence signatures in silico so one of the immediate goals that we 01:18:01.460 --> 01:18:08.420 became interested in during the outbreak was to develop a platform of viruses that captured the 01:18:08.500 --> 01:18:14.340 heterogeneous that exists within the family of viruses and to do that we identified sort of 01:18:14.340 --> 01:18:19.140 representative strains for example middle phase early phase isolate or bony we had in the lab 01:18:19.140 --> 01:18:26.740 middle phase isolate with cohk w1 kzo2 with early phase isolate animal isolates like it's from sivets 01:18:26.740 --> 01:18:34.100 fc 16 and hc fc 6103 a raccoon dog isolate which is shown down here the sporadic 2004 human case 01:18:35.060 --> 01:18:41.380 and a couple of other viruses sequences were identified as good candidates that would capture 01:18:41.380 --> 01:18:48.180 all the diversity that had been identified within the sequence pool this shows the sequence that's 01:18:48.180 --> 01:18:54.100 the sequence variation within the spike like a protein if it's a blue box amino acid that's an 01:18:54.100 --> 01:18:59.780 animal associated residue early phase changes are shown here in yellow middle phase changes 01:18:59.780 --> 01:19:05.860 shown in orange and late phase changes shown in red in addition in 2004 there are a variety 01:19:05.860 --> 01:19:10.580 of other animal strains of sporadic human cases were identified that had additional variations 01:19:10.580 --> 01:19:16.820 shown here in white and it's interesting as much i i hope that everybody was paying attention right 01:19:16.820 --> 01:19:25.620 what i put up here before you were paying attention right um this paper is actually 01:19:26.580 --> 01:19:32.740 this paper this paper right here 01:19:36.740 --> 01:19:42.660 broad spectrum coronavirus antiviral drug discovery by allicin totura and cena bivari 01:19:42.660 --> 01:19:48.420 written in 2019 allicin totura was a postdoc of ralph veric 01:19:48.420 --> 01:19:55.620 so she's at us amrit 01:19:59.140 --> 01:20:01.620 you don't think she knows how to make infectious clones 01:20:03.620 --> 01:20:07.300 you don't think everybody that went through ralph veric's lab knows how to make infectious 01:20:07.300 --> 01:20:13.860 clones are you kidding me where did these people go off to they're not working at starbucks or 01:20:13.860 --> 01:20:16.980 something like that you think 01:20:20.500 --> 01:20:25.460 this paper is not accidental that it talks about a virus getting out of china and needing to use 01:20:25.460 --> 01:20:32.900 remdesivir as an antiviral for it it's not for nothing that we're defending the proof reading 01:20:32.900 --> 01:20:41.460 ability of exo x one gene or or our exo and gene uh the the ribonuclea we're not we're not 01:20:41.460 --> 01:20:49.700 defending these these things for nothing they're all related all these people all their nonsense 01:20:49.700 --> 01:20:58.740 it's all related don't you remember the the the domain program run by ditra 01:21:00.340 --> 01:21:06.820 run by robert malone who supposedly made an x-ray crystallography computer model 01:21:06.900 --> 01:21:15.060 of the three cl protease of the virus and then interface that computer model with all known 01:21:15.060 --> 01:21:20.500 pharmaceuticals with a volunteer team of researchers in a matter of weeks to identify 01:21:20.500 --> 01:21:29.300 femotidine and remdesivir as two usefully useful antivirals 01:21:37.460 --> 01:21:40.020 and cena bivari knew about it a year before that 01:21:41.700 --> 01:21:45.620 and ralph veric and mark dennison knew about it a few years before that 01:21:46.580 --> 01:21:51.220 for zika or for for Ebola for whatever else they were trying it on 01:21:56.500 --> 01:21:58.740 because they're all RNA viruses right 01:22:00.980 --> 01:22:06.660 don't you see the pattern here there is a mythology that has been laid down there is a 01:22:06.660 --> 01:22:08.500 mythology that is being defended 01:22:08.900 --> 01:22:20.580 this paper got in bobbie's book this paper that is in bobbie's book and actually actually 01:22:20.580 --> 01:22:26.500 right uh uh robert malone sold bobbie's book today on his sub stack which is pretty badass i like 01:22:26.500 --> 01:22:33.460 that a lot maybe that was part of the deal if you get that you stop that kooie from talking about 01:22:33.540 --> 01:22:40.580 me on his stream i'll i'll sell bobbie's book on my sub stack maybe that's what it is 01:22:41.780 --> 01:22:46.260 it's a pretty simple deal right you know robert malone's got a million subscribers if he tweets 01:22:46.260 --> 01:22:49.860 out a book probably a hundred thousand people buy it or ten thousand people buy it 01:22:52.660 --> 01:22:57.540 he better sub stack the book he wrote a wrote a review on the back cover 01:22:58.420 --> 01:23:06.900 i'd say buy a copy i'd say buy a copy and we'll meet and i'll autograph it for you 01:23:07.860 --> 01:23:12.820 um there's some bad stuff in there there's some good stuff in there it's probably the best 01:23:12.820 --> 01:23:14.820 document of all the nonsense people said 01:23:18.580 --> 01:23:22.980 the person who showed me this paper the person who showed me this paper his Mark 01:23:23.540 --> 01:23:29.780 Housatonic Mark Kulacz it wasn't shown to me by by Kevin McCairn it wasn't shown to be by 01:23:29.780 --> 01:23:36.580 Paul Cottrell it wasn't shown to me by George Webb wasn't shown to me by Kevin McKernan or Stephanie 01:23:36.580 --> 01:23:44.580 Sinev or Steve Kirsch or Robert Malone the only person that's ever mentioned this paper to me 01:23:44.580 --> 01:23:48.740 ever shown it to me ever found it and said holy crap dude did you read this 01:23:49.300 --> 01:23:51.780 this is Mark Kulacz 01:23:55.780 --> 01:24:00.900 i just can't stress to you how all of these things line up in a way that that can't be 01:24:00.900 --> 01:24:06.340 discounted anymore all of these people all of their behavior line up in a way that cannot 01:24:06.340 --> 01:24:11.300 be discounted anymore even if we find out that there's a huge coronavirus background that 01:24:11.300 --> 01:24:18.180 coronaviruses can transmit for months even if we find out that clones can transmit for months 01:24:19.620 --> 01:24:26.580 the ultimate conclusion is is that we have been lied to by a group of medlers that sustained 01:24:26.580 --> 01:24:34.980 a worst-case scenario sustained a confusion and uncertainty and a doubt that allowed the world 01:24:34.980 --> 01:24:41.620 to be transfected and only after the world was transfected that these people do it about face 01:24:41.620 --> 01:24:47.940 and pretend like they always knew and the list of these people is long but it doesn't include 01:24:47.940 --> 01:24:55.460 Ralph Baric the list of people who lied to us about the worst-case scenario at the beginning 01:24:55.460 --> 01:25:02.020 of the pandemic only to pivot after our kids had lost two years of school after our kids have 01:25:02.020 --> 01:25:09.220 been traumatized with masks after we had been locked down for a year and a half that's not Ralph Barrick 01:25:09.220 --> 01:25:21.380 case you can't tell i'm trying to get Ralph Baric to come and interview on Gigaohm 01:25:21.380 --> 01:25:27.220 Biological wouldn't that be badass much of the variation actually falls within residues or regions 01:25:27.220 --> 01:25:33.620 that neutralizing epitopes were identified and in fact although the number of residues that are 01:25:33.620 --> 01:25:38.420 changed aren't that great you can actually reduce neutralization titers by about 20 fold 01:25:39.460 --> 01:25:47.300 with some of these variants now importantly Farzan's group showed that the two key changes 01:25:47.300 --> 01:25:52.500 during the epidemic where this life seemed to asparaging change at 4.79 and Assyrian 3 01:25:52.500 --> 01:26:00.340 change at 4.87 that drove animal adaptation to human strain and so keep that in mind so 01:26:01.060 --> 01:26:10.340 without access to this pool of strains we decided to synthesize basically a portfolio of 01:26:11.060 --> 01:26:16.980 spike like a protein genes of about 4k each and then use a molecular clone for the 01:26:16.980 --> 01:26:22.100 urbani epidemic strain that we had built in the lab basically replacing the urbani spike 01:26:22.100 --> 01:26:26.020 one at a time with these various spike like a protein from different phases of the epidemic 01:26:26.820 --> 01:26:32.580 now all of these viruses were viable except for the sc-16 variant this actually can't recognize 01:26:32.580 --> 01:26:38.100 the humanase receptor so it didn't grow until we made mouth cells that consistently expressed 01:26:38.100 --> 01:26:42.980 the civetase to receptor and once we did that this virus to grow well before we had those cells 01:26:43.620 --> 01:26:50.260 since Farzan had identified this 4.79 changes the key residues so be careful there now they 01:26:50.260 --> 01:26:55.620 just admitted that they use mouth cells which have been genetically altered to express the 01:26:55.620 --> 01:27:02.900 civetase receptor now i can't stress enough we had this paper come out that everybody's been 01:27:02.900 --> 01:27:07.460 yelling about it i'm probably gonna have to do a show about it in the next couple days again 01:27:08.260 --> 01:27:14.980 about this paper that um you know i should do that video tomorrow that's what i'm gonna do 01:27:14.980 --> 01:27:24.980 tomorrow i'm gonna do i found this video with mccernan and um christine uh parks christina parks 01:27:25.860 --> 01:27:35.140 and um John Beaudoin and i've i've been meeting to do John Beaudoin um on HighWire and so I should 01:27:35.140 --> 01:27:41.860 do John Beaudoin on HighWire this week and then I'll do this this multiple person stream that 01:27:41.940 --> 01:27:46.740 happened a few days a few a week ago or so with McKernan and parks and Beaudoin that'll be good 01:27:46.740 --> 01:27:52.900 that's two good good study halls for this week um so what he's talking about here now is making a 01:27:52.900 --> 01:27:57.940 number of different clones where he's swapping the spike out um this is of course supposed to get 01:27:57.940 --> 01:28:03.300 us really excited about making chimeric viruses but here again he's gonna make clones of all these 01:28:03.300 --> 01:28:09.700 things so if there's a danger the danger is is that he's making clones if he wasn't making clones he 01:28:09.700 --> 01:28:16.740 couldn't even study these things the regular virus of these things is not attainable won't grow 01:28:16.740 --> 01:28:21.940 and even if you could grow it the titers would be so low it would be almost unsequitable 01:28:23.300 --> 01:28:30.980 that's the problem with these guys right and so don't let the the nonsense fool you 01:28:30.980 --> 01:28:38.420 infectious clones are the bedrock methodology of RNA virology change especially build a recombinant 01:28:38.420 --> 01:28:44.180 virus with that change and that virus could be cultured uh interestingly enough two of these 01:28:44.180 --> 01:28:50.340 the gz-o-2 and the uh hcse-61 of three viruses actually caused lethal infection in aged animals 01:28:50.340 --> 01:28:56.020 they produced an ARDS like disease saw SARS caused ARDS in humans or predominantly in age 01:28:56.020 --> 01:29:01.300 populations uh that's acute respiratory distress syndrome it's a clinically devastating end stage 01:29:01.300 --> 01:29:06.420 lung disease with about a 50 mortality rate so these are actually some of the first real good 01:29:06.420 --> 01:29:13.300 animal models for SARS infection and so remember what I said the other day again you have to 01:29:14.100 --> 01:29:19.380 take everything with a huge grain of salt if it's an animal model especially a genetically modified 01:29:19.380 --> 01:29:27.700 one then the expression of the protein that the virus interacts with could be so over or so 01:29:27.780 --> 01:29:39.140 inappropriate or any other number of of wrong that when exposed to these exosome collections 01:29:39.140 --> 01:29:44.740 these animals produce a really robust immune response or a really robust infection because 01:29:45.300 --> 01:29:49.780 these signals go everywhere so if you believe the biology that they need a receptor and that 01:29:49.780 --> 01:29:54.420 the receptor has to be somewhere and if the receptor isn't there then the virus can't hurt you 01:29:54.980 --> 01:30:01.700 if you believe this then these animal models are also very scary because you could imagine 01:30:01.700 --> 01:30:07.300 a scenario whereby genetically altering the animal you create an animal that is hyper 01:30:07.940 --> 01:30:14.740 susceptible to this manipulation hyper susceptible to the exposure to these signals 01:30:15.700 --> 01:30:22.740 so if you believe viruses work the way that these people say they do and that they need receptors 01:30:22.820 --> 01:30:27.940 and then you alter the expression of those receptors or even worse you homogenize them 01:30:27.940 --> 01:30:34.180 across the whole animal then their susceptibility to exposure to these things could be very different 01:30:34.180 --> 01:30:40.180 and that that could be great if you're writing grant applications that need an animal model that's 01:30:40.180 --> 01:30:46.900 very repeatable and very robust and so don't underestimate the stuff that we talked about with 01:30:47.460 --> 01:30:53.620 with Wolfgang Wodock keeps coming up here where you have academic biologists that are essentially 01:30:53.620 --> 01:31:01.060 academy-gicians they've been they become so good at doing experiments that are fundable 01:31:02.020 --> 01:31:08.980 asking questions that are fundable that they have stopped asking useful questions that they have 01:31:08.980 --> 01:31:14.900 stopped asking questions that actually move the ball forward but they're now asking sort of circular 01:31:14.980 --> 01:31:22.580 questions they don't want to make any progress they want to make the illusion of motion right 01:31:24.420 --> 01:31:29.140 and so in this scenario i don't think that's what what Ralph barracks doing at all i think he's 01:31:29.140 --> 01:31:35.460 actually making forward progress there are these RNA signals which have been very difficult to study 01:31:35.460 --> 01:31:41.940 in the wild because they're almost unculturable there's no infectious material available it's so 01:31:41.940 --> 01:31:47.300 rare and if you take this infectious material and try to culture it you often get very low to 01:31:47.300 --> 01:31:58.260 no tighter so infectious clone overcomes that Ralph barrack is a genius now um these viruses 01:31:58.260 --> 01:32:03.780 replicate of the human epidemic strains replicate efficiently in human airway epithelial cultures 01:32:03.780 --> 01:32:08.500 these are cultures that are derived from cells lining the trachea of transplant patients you can 01:32:08.580 --> 01:32:13.780 actually culture them on liquid air interfaces and they take on the architecture of the human airway 01:32:13.780 --> 01:32:20.420 stars coronavirus all the epidemic strains actually like to infect ciliated cells and these epidemic 01:32:20.420 --> 01:32:26.900 strains replicate very efficiently the animal strains however do not and this is just some 01:32:26.900 --> 01:32:33.540 fluorescent microscope images showing the cilia of the ciliated cells with an expression of the 01:32:33.540 --> 01:32:39.220 sars nucleic acid protein from an epidemic strain on the ciliated cells and the zoonotic 01:32:39.220 --> 01:32:47.140 strains se 16 in this and hdse 6103 and replicate however if you passage these viruses on human 01:32:47.140 --> 01:32:52.660 airway cells and culture you can actually rapidly select out variants that can replicate efficiently 01:32:52.660 --> 01:32:59.540 in human airways those viruses actually do not contain the mutations that would be had been 01:32:59.620 --> 01:33:05.780 predicted by mike farson as being which were clearly responsible for the 2002-2003 epidemic 01:33:06.340 --> 01:33:14.100 rather we saw changes um at positions 442 and 472 and 479 and so this is an interesting 01:33:14.100 --> 01:33:23.300 presentation of the data where it seems like this single trial of putting a clone on a epithelial 01:33:23.300 --> 01:33:30.100 cell culture a differentiated epithelial cell culture and then seeing sequence changes in the 01:33:30.740 --> 01:33:37.860 consensus sequence he seems to talk about it like okay so we put the car on the road and it drove 01:33:37.860 --> 01:33:42.180 down the road and then that's how it works and so if we did it again that's what would happen 01:33:43.380 --> 01:33:49.300 and i don't think that that's really how virology works i think if he did this again he would get 01:33:49.300 --> 01:33:57.300 different results here it might be a similar kind of selection process by which non-infectious 01:33:57.300 --> 01:34:04.580 particles versus infectious particles are selected and and can be seen in in in subsequent passages 01:34:04.580 --> 01:34:10.660 or detected in subsequent passages but that selection process is purely based on function 01:34:10.660 --> 01:34:18.020 then it's not going to be based on fitness nothing is is fitness unless it's number of copies right 01:34:18.020 --> 01:34:23.300 so then we're talking about a whole different thing and we're really only doing a selection 01:34:23.300 --> 01:34:28.340 process we're taking the whole supernatant here there's there's no like you know 01:34:29.540 --> 01:34:34.740 race to see who can make more copies of themselves we're not selecting based on fidelity or anything 01:34:34.740 --> 01:34:38.980 like that in fact we have the argument that there has to be mutation rate otherwise there would be 01:34:39.060 --> 01:34:46.660 a collapse so it's interesting um that's it's that's it mediated the cross species transmission 01:34:46.660 --> 01:34:52.500 event so in reality we've done this a couple of times there's actually several pathways by which 01:34:52.500 --> 01:34:57.940 the zoonautics are it could actually adapt and recognize the humanase receptor what's interesting 01:34:57.940 --> 01:35:04.340 is that the epidemic strains actually efficiently use both the humanase receptor and the civetase 01:35:04.420 --> 01:35:10.260 receptor when we in vitro select for human adapted strains on human airway culture 01:35:11.140 --> 01:35:16.820 they actually lose the ability to recognize the civic receptor so what this data suggests 01:35:16.820 --> 01:35:22.900 actually is that ours had existed in a transmission cycle between humans and civets actually probably 01:35:22.900 --> 01:35:28.500 for several several years prior to the 2002-2003 epidemic allowing the virus to 01:35:28.580 --> 01:35:34.900 regain the capacity to recognize both receptors okay so those were the easy ones to do 01:35:35.860 --> 01:35:41.700 oh with an application um so now that we had a large panel of our of uh variant viruses we could 01:35:41.700 --> 01:35:46.500 use those for therapeutic testing over vaccines in this case i'm going to show you an example of 01:35:46.500 --> 01:35:52.420 about 30 human monoclonal antibodies that were derived by Antonio Lanzavecchia if you take those 01:35:52.420 --> 01:35:56.820 30 monoclonals and test their ability to neutralize all the strains that we've made in the laboratory 01:35:56.820 --> 01:36:03.220 you find some that only neutralize urbani some that neutralize all human strains some that neutralize 01:36:03.220 --> 01:36:08.820 some civets but not all the animal strain but you do end up with four antibodies that neutralize 01:36:08.820 --> 01:36:14.340 all strains we have in our portfolio including the ones that were in vitro adapted on human 01:36:14.340 --> 01:36:22.260 airway culture importantly if you select for escape using these antibodies these antibodies 01:36:22.260 --> 01:36:26.420 select for changes in different locations of the receptor binding domain 01:36:26.420 --> 01:36:32.020 and in fact three of them actually don't overlap yes 109 the 230 and the 227 01:36:32.020 --> 01:36:37.380 don't overlap and so these represent good therapeutic cocktails that would capture most of the 01:36:37.380 --> 01:36:43.460 diversity that exists in SARS therapeutic cocktail what an interesting story we can tell there so 01:36:44.260 --> 01:36:51.380 there's a guy by the name of plumber who used to work on AIDS in Canada who died at the beginning 01:36:51.460 --> 01:37:03.140 of the pandemic he had a postdoc I think she was Chinese and she had a three antibody cocktail 01:37:03.140 --> 01:37:13.620 called Z map that was trialed against remdesivir and was actually being trialed against remdesivir 01:37:13.620 --> 01:37:20.100 probably at us amrit right before the pandemic and then it was shut down 01:37:21.700 --> 01:37:30.020 so the trial comparing remdesivir with Z map was stopped this is all Kevin sorry this is all 01:37:30.020 --> 01:37:39.140 marcoolax work here that I'm reciting from memory so we have an AIDS expert dying I think he fell 01:37:39.140 --> 01:37:46.980 on the sidewalk or something crazy like that and we have this spy Chinese lady who supposedly 01:37:48.100 --> 01:37:53.700 took some samples back to China and so she's a spy she she got put away but actually she was 01:37:54.260 --> 01:38:03.540 like on the cover of magazines and stuff is being the star of of of biology one year for Z map it's 01:38:03.540 --> 01:38:11.460 really cool because that was a three antibody cocktail this is also another proposed three 01:38:11.460 --> 01:38:21.220 antibody cocktail as a as a pan by an antiviral that's pretty that's pretty cool I I'm starting 01:38:21.220 --> 01:38:28.020 to see Ralph Barrett as a good guy it's weird and probably work in potential patients that would 01:38:28.100 --> 01:38:33.140 be infected during future epidemics and these also actually protect young and aged animals from 01:38:33.140 --> 01:38:40.260 lethal infection that was a paper published by Barry Rock in a couple papers now the civet 01:38:40.260 --> 01:38:46.420 and the raccoon dog strains were the easy ones the true reservoir for SARS is within bat population 01:38:47.460 --> 01:38:52.980 so if you look at a phylogenetic tree the human strains are shown here in red the civet strains 01:38:52.980 --> 01:38:57.540 raccoon dog strains are shown in purple but the real variation is within the bat strain 01:38:58.580 --> 01:39:02.340 um the reservoir it's thought that these were probably the reservoir for the 01:39:03.140 --> 01:39:09.220 emergence of the virus now these are about 80 to 90 percent identical to SARS they can't be 01:39:09.220 --> 01:39:14.100 cultured and they exist to sequence signature signatures in silico there's also extensive 01:39:14.100 --> 01:39:20.020 variation see they can't be cultured they exist as sequence signatures in silico well that just 01:39:20.020 --> 01:39:26.900 means they're stored on a computer they're keyboard viruses and so clones are really really important 01:39:26.900 --> 01:39:33.940 because you can take a virus that you've only detected in the wild as a sequence and create 01:39:33.940 --> 01:39:41.060 large quantities of that RNA and electropyrated into cells and get those cells to run it through 01:39:41.060 --> 01:39:49.460 their ribosomes and package it the only thing you can't do sometimes is passage it 01:39:49.940 --> 01:39:57.940 but the cells can be made to take the RNA up they can be it can be electropyrated in 01:40:00.020 --> 01:40:05.540 you can use lip effectamine to get it in and then those cells will read the RNA and those 01:40:05.540 --> 01:40:09.780 proteins will produce and the sub genomic RNAs will be produced and everything will be packaged 01:40:09.860 --> 01:40:20.740 up and then you'll get this this effect this is cool because this is really underscoring 01:40:22.580 --> 01:40:29.700 that downplaying the clones is just it's absurd it's absurd it is a bedrock methodology of RNA 01:40:29.700 --> 01:40:34.980 virology through the replicase and elsewhere in the genome so when we decided we were going to 01:40:34.980 --> 01:40:41.620 synthetically resurrect the entire virus now before we started it's important to note this is a 01:40:44.340 --> 01:40:51.220 the cryoem reconstruction of the SARS glycoprotein spike notice that there are three receptor binding 01:40:51.220 --> 01:40:57.300 domains shown here in with bubbles that actually engage the ACE2 receptor if you look at the sequence 01:40:57.300 --> 01:41:03.860 the receptor binding domain shown here in yellow there's a large amount of sequence variation 01:41:03.860 --> 01:41:09.060 especially within the contact interface residues that engage the human ACE receptor 01:41:09.060 --> 01:41:13.620 and in fact there's only four of 13 contact interface residues that are retained in these 01:41:13.620 --> 01:41:18.580 fat strains but we don't actually think it's going to use the human ACE receptor to get into cells 01:41:19.300 --> 01:41:24.580 in fact we think we're going to have a tough time culturing the virus now it's important to note 01:41:24.580 --> 01:41:31.700 that the SARS RBD that is was identified has been proposed by a couple of groups that it may have 01:41:31.700 --> 01:41:37.220 been introduced by recombination processes from unknown strains that haven't yet been identified 01:41:37.220 --> 01:41:42.820 and that that led to the initial cross-species transmission events now to get back to the 01:41:42.820 --> 01:41:48.900 issue of in silico sequences they actually represent hypothetical viruses most synthetic viruses that 01:41:48.900 --> 01:41:55.540 have been resurrected to this point actually we knew that the sequence was infectious in this case 01:41:55.540 --> 01:42:01.460 we actually don't know which of the sequences in GenBank were infectious if any so to do that 01:42:01.460 --> 01:42:05.860 with an error rate in GenBank ranging from about one to five hundred to one to ten thousand depending 01:42:05.860 --> 01:42:12.100 on the sequence we had to do extensive bioinformatic analysis to identify what we thought was the likely 01:42:12.100 --> 01:42:18.020 consensus sequence well that sounds like that sequencing is a lot more hairy than i thought it was 01:42:18.740 --> 01:42:22.980 I just thought sequences were sequences you get one it just kind of spits it out 01:42:22.980 --> 01:42:26.900 beep beep beep beep beep beep beep beep beep beep beep beep beep beep beep beep beep beep beep 01:42:26.900 --> 01:42:32.660 then you just kind of read it off right now we're getting back to the Vincent Rancin yellow 01:42:32.660 --> 01:42:41.940 explanation of consensus sequence the the more traditional Vincent Rancin yellow will say that 01:42:41.940 --> 01:42:50.820 the consensus sequence might not even actually exist but Kevin McCurnan says that there are 15 01:42:50.820 --> 01:42:57.380 million SARS-CoV-2 genomes on on gents said they're all real they're all independent they all come 01:42:57.380 --> 01:43:03.140 from different people so they have to be real what's Ralph Barrick talking about here then 01:43:03.940 --> 01:43:16.500 I see a battleship size incongruency between pre-COVID virology and post-COVID virology 01:43:19.860 --> 01:43:25.300 I really do I really think we're on to something here I think with a couple weeks of work 01:43:25.940 --> 01:43:34.260 this is going to be over and we're going to be able to you know incorporate all of these 01:43:34.260 --> 01:43:40.500 various factions into a common explanation for why a lot of people were mistaken 01:43:41.460 --> 01:43:47.700 why a lot of people were fooled why you could be taken all the way by no virus why you could 01:43:47.700 --> 01:43:52.900 be taken all the way by lab leak why you could be taken all the way by my market zoneosis and 01:43:52.980 --> 01:43:58.660 why you could be taken all the way by by any of these other in-betweens including 01:44:00.100 --> 01:44:01.700 including iatrogenic murder 01:44:04.420 --> 01:44:11.460 but in the end there's only one explanation that fits all of them that lets the most people on 01:44:11.460 --> 01:44:22.660 earth be right and I think that's why everybody's so angry because the explanation that we are being 01:44:22.660 --> 01:44:28.900 fooled into believing it's Ralph Barrick and Tony Fauci and eco-health alliance 01:44:30.420 --> 01:44:39.940 instead of understanding that this technology was actually available available it was available 01:44:40.900 --> 01:44:41.860 to us Amrit 01:44:44.420 --> 01:44:50.020 probably for a couple decades right ever since they first did it with the polio virus 01:44:53.540 --> 01:44:57.860 we have to wake up and apologize to our kids sooner or later we might as well do it now 01:44:58.660 --> 01:45:03.380 none of the strains that we actually saw we thought was completely correct some of them had 01:45:03.380 --> 01:45:08.980 deletions in the sequence of the five prime ends that we had to make some educated guess 01:45:10.180 --> 01:45:14.900 the basic approach that we build coronavirus is using our molecular clone is shown here 01:45:14.900 --> 01:45:21.860 with the SARS clone shown in blue is the clone is broken into six five kv about five kv thesis 01:45:22.660 --> 01:45:29.620 each piece is flanked by bagel one restriction endonucleate sites these are class two S restriction 01:45:29.620 --> 01:45:35.060 enzymes that recognize a palindromic sequence and so this is asymmetric and this is almost 01:45:35.060 --> 01:45:40.020 certainly one of the enzymes that was mentioned in the foyer request that was recently released 01:45:41.300 --> 01:45:50.580 and so the release of this enzyme or related enzymes as a list in an email or a supplementary 01:45:50.580 --> 01:45:56.580 document or something like that is now being misconstrued as evidence that the diffuse proposal 01:45:56.580 --> 01:46:01.060 is real and that they were making clones and that they were spraying them into bad caves and they 01:46:01.060 --> 01:46:08.980 put fear and cleavage sites at the joint between S1 and S2 yada yada yada so most likely that's 01:46:08.980 --> 01:46:15.780 why we're being fed this video is so that we get to this point right here and we see how clones 01:46:15.860 --> 01:46:24.420 are assembled and now i'm sure he's about to tell us that this right here represents a S2 S1 S2 01:46:26.340 --> 01:46:32.420 clone junction there so that they can substitute the S1 from other coronavirus is right in there 01:46:32.420 --> 01:46:40.260 let's listen this allows since these ends are asymmetric they actually will not concatomerize 01:46:40.260 --> 01:46:44.580 like classic sticky ends left by restriction enzymes but rather they become directional 01:46:45.220 --> 01:46:50.660 so if you end clone A with a bagel site that leaves one three nucleotide overhags and the 01:46:50.660 --> 01:46:55.060 five prime end of the B fragment with the complementary three nucleotide overhags they 01:46:55.060 --> 01:46:59.780 totally get it directly you change different bagel sites at each piece it's midnight 01:47:00.740 --> 01:47:07.940 assemble up into 30k B chromosomes like tinker toys like tinker toys like tinker toys 01:47:07.940 --> 01:47:13.300 batching home that we made that's actually a good way to date somebody like do you know 01:47:13.300 --> 01:47:19.140 what tinker toys man i had tinker toys you know i had lots of them like a lot of them i had a 01:47:19.140 --> 01:47:25.300 lot of tinker toys i think i had like three of the cylindrical containers i could go down 01:47:25.300 --> 01:47:30.740 with tinker toys hey by the way it's uh past midnight so technically speaking i just turned 01:47:30.740 --> 01:47:39.860 52 years old yeah 52 52 years old how's that for old using blue heron and biobasic basically 01:47:39.860 --> 01:47:45.060 is interchangeable with the urbanic clone the only real difference was that we broke the F 01:47:45.060 --> 01:47:50.180 fragment into two pieces so that we could play with the receptor binding domain easily if this 01:47:50.180 --> 01:47:56.020 thing didn't turn out to be infectious and in fact we made this clone we built the fool on cDNA we 01:47:56.020 --> 01:48:00.660 drove transcripts electroprated that in the cells and we can see evidence of replication by 01:48:00.660 --> 01:48:05.540 the synthesis of sub-genomic messenger RNA but we couldn't culture the virus and we couldn't pass 01:48:05.540 --> 01:48:10.420 it from cell to cell ah we couldn't culture the virus we couldn't passage it from cell to cell so 01:48:10.420 --> 01:48:17.460 they could see that the RNA was being sorted into sub-genomic RNAs but they couldn't passage it from 01:48:17.460 --> 01:48:23.620 cell to cell does that mean it was not getting packaged or does that mean that it wasn't binding 01:48:23.620 --> 01:48:28.980 in the next passage i think it wasn't binding in the next passage is the right explanation 01:48:28.980 --> 01:48:35.940 which means that you could make virus from these clones you just have to have a receptive cell type 01:48:35.940 --> 01:48:40.900 on the other end that's going to do it so they transfected listen carefully they transfected 01:48:41.540 --> 01:48:49.460 this clone into that cell culture and it packaged virus for them you know it packaged virus because 01:48:49.460 --> 01:48:54.420 it made the sub-genomic RNAs those were translated into the many many many copies of the proteins 01:48:54.420 --> 01:48:59.860 and all this stuff happened it's just that when they took the supernatant off of that cell culture 01:48:59.860 --> 01:49:06.180 and tried to put it on the next cell culture it didn't propagate and that has to do with the spike 01:49:06.180 --> 01:49:11.860 protein receptor binding domain compatibility with the cell culture that you're using according to 01:49:11.860 --> 01:49:18.020 their methodology explanation so let's listen to that again because i want you to hear what he did 01:49:18.100 --> 01:49:22.900 he electroporated it into a cell culture but couldn't get it to jump from cell culture to cell 01:49:22.900 --> 01:49:30.820 culture which is what culturing is what passaging is clearly there was RNA the cell and we could 01:49:30.820 --> 01:49:36.100 clone we built the full-on cDNA we drove transcripts electroprated that in the cell and we can see 01:49:36.100 --> 01:49:40.740 evidence of replication by the synthesis of sub-genomic messenger RNA but we couldn't 01:49:40.740 --> 01:49:45.380 culture the virus and we couldn't pass it from cell to cell sub-genomic electroprated that and in fact 01:49:45.380 --> 01:49:50.660 we made this clone we built the full-on cDNA we drove transcripts electroprated that in the cell 01:49:50.660 --> 01:49:55.380 and we can see evidence of replication by the synthesis of sub-genomic messenger RNA 01:49:55.380 --> 01:49:59.780 but we couldn't culture the virus and we couldn't pass it from cell to cell so clearly there was 01:49:59.780 --> 01:50:08.740 probably a defect in entry to solve that problem we use literature data that has suggested that 01:50:08.740 --> 01:50:14.020 RBD domains of coronaviruses may be interchangeable between species so we took the human 01:50:14.900 --> 01:50:20.580 the urbani epidemic receptor-bonding domain that's 210 amino acids and went 01:50:21.620 --> 01:50:28.260 and dropped that into the bat genome backbone producing a chimera with the receptor-bonding domain 01:50:28.260 --> 01:50:33.940 driven from the epidemic strains now when we built that clone drove transcripts and electroprated 01:50:33.940 --> 01:50:39.140 that in the cells we got a virus that could replicate quite efficiently this is just some 01:50:39.140 --> 01:50:46.020 growth curve data showing I think the black box is the urbani wild type and the white circles are 01:50:46.020 --> 01:50:52.980 open open symbols are the bat viruses you can see they replicate exactly as good as the urbani 01:50:52.980 --> 01:50:58.900 epidemic strain at low and high multiplicities of infection they recognize the human ACE2 receptor 01:50:58.900 --> 01:51:04.420 and dbt cells at low and high multiplicity infections and grow just like the epidemic strains 01:51:04.420 --> 01:51:10.020 and they also retain the ability to use the CIVID ACE2 receptor just like the epidemic strains 01:51:10.020 --> 01:51:16.500 low and high multiplicity they also are capable of infecting human airway epithelial cultures 01:51:16.500 --> 01:51:21.380 and targeting ciliated cells just like the epidemic strain and they grow to similar titer 01:51:23.700 --> 01:51:28.740 now if you make aniseera just against the bat spike like a protein 01:51:29.620 --> 01:51:36.500 and use that aniseera to target the SARS wild type virus it will not neutralize that virus 01:51:37.220 --> 01:51:43.140 so clearly aniseera and vaccines derived against epidemic strains are not going to protect against 01:51:43.140 --> 01:51:49.540 the bat strains to use that same sera against the RBD chimeric virus you can neutralize it 01:51:49.540 --> 01:51:55.220 indicating that there are neutralizing episodes that resist that exist outside the RBDs and if 01:51:55.220 --> 01:52:00.340 you take the human monoclonal to target the epidemic strain RBDs you can neutralize these 01:52:00.340 --> 01:52:05.700 virus with quite efficient so one thing that I think we all need to learn is exactly what neutralized 01:52:05.700 --> 01:52:13.460 means in this context because I kind of was under the naive operation or operating under the naive 01:52:13.460 --> 01:52:20.580 assumption that that neutralized means to bind to the receptor binding domains so that the receptor 01:52:20.580 --> 01:52:24.580 binding domain is not able to interact with the receptor and therefore you don't get 01:52:25.540 --> 01:52:30.420 infection so I thought that's what neutralizing was but I think neutralizing has more to do with 01:52:30.420 --> 01:52:38.900 like precipitation and if you get enough antibodies stuck onto you then you come out of out of solution 01:52:38.900 --> 01:52:46.020 and I'm wondering if it's something like this too like uh aggregation or something like that 01:52:46.020 --> 01:52:51.460 that causes neutralization and precipitation so I need to learn that a little bit better because 01:52:51.460 --> 01:52:57.860 I do know that that people like Mark Bailey and and uh Steven Lanka have been working on trying to 01:52:57.860 --> 01:53:02.900 understand what it is that virologists are meaning with that and I do think that that is something 01:53:02.900 --> 01:53:11.220 that we should be masters of here as well so I apologize for that so in summary um certainly 01:53:11.220 --> 01:53:15.780 synthetic genomics and reverse genetics this can be a platform to recover uncultivatable 01:53:15.780 --> 01:53:20.580 zoonotic precursors can be used to synthesize this is actually the largest synthetic virus 01:53:20.580 --> 01:53:27.540 to date it's going to be a short record but it is a record um and you can use it now to identify 01:53:27.540 --> 01:53:33.860 a broad spectrum antiviral from the vaccine clearly the RBDs are interchangeable uh this is with an 01:53:33.860 --> 01:53:40.980 89 percent amino acid identity within the spike it's the minimal domain required to host shift 01:53:40.980 --> 01:53:46.420 coronaviruses what is the phylogenetic limits to RBD interchange we actually don't know 01:53:47.620 --> 01:53:52.580 clearly the human monoclonals and vaccines targeting the SARS RBD would provide protective 01:53:52.580 --> 01:53:56.740 immunity against natural isolates that emerge that's not my sound the overall process is in 01:53:56.740 --> 01:54:04.500 future or unfortunately by deliberate design so now I want to move to the next part of the 01:54:04.500 --> 01:54:09.700 talk which is synthetic the optimization scheme to attenuate SARS coronavirus pathogenesis 01:54:10.580 --> 01:54:16.420 this is the maturation pathway okay so after as he explains the maturation pathway I want you to 01:54:16.980 --> 01:54:22.340 imagine that we're trying to solve the puzzle of the infectious cycle as well and then I also 01:54:22.340 --> 01:54:30.820 want you to listen carefully to the amount of effort time thought and ideas that are going into 01:54:31.620 --> 01:54:42.340 the attenuation of coronaviruses and a general strategy to attenuate all coronaviruses specifically 01:54:42.340 --> 01:54:50.580 in the context of recombination and so he's keenly aware that there's a problem with the 01:54:50.580 --> 01:54:56.100 oral poliovirus vaccine that when you give the oral poliovirus vaccine it can mix with other 01:54:56.180 --> 01:55:03.460 polioviruses in the gut of people and then become active polio again and he's at least 01:55:03.460 --> 01:55:09.300 theoretically aware of the possibility that should you make a de attenuated coronavirus like say 01:55:10.100 --> 01:55:17.300 take out some genes then it would be very easy for that clone that you're using as an attenuated 01:55:17.300 --> 01:55:24.500 virus to attain that gene to acquire that gene from a co-infection and so what he's going to explain 01:55:24.500 --> 01:55:32.580 to you is how these translational regulatory sequences can be altered in such a way to prevent 01:55:32.580 --> 01:55:38.500 recombination with wild type viruses that's his strategy I think it's a pretty cool idea 01:55:39.300 --> 01:55:46.340 it suggests that these self-replicating RNAs are are some kind of a real biological phenomenon 01:55:47.220 --> 01:55:52.980 and it suggests that making infectious clones of them is a really handy way of exploring the 01:55:52.980 --> 01:56:01.140 variation within them that is otherwise un-intractable by laboratory means so here we go again I 01:56:01.140 --> 01:56:07.540 apologize that this is such a fuzzy thing but I will try to help you follow along way for how 01:56:07.540 --> 01:56:12.500 far it gets out of the cell the nuclei caps is the line underneath an intermediate compartment 01:56:12.500 --> 01:56:19.140 in the ruffiardology with a M glycoprotein and the E-protein are expressed and then these the M 01:56:19.140 --> 01:56:24.900 and the E-protein actually drive the assembly of virion which then mature in the Golgi and then 01:56:24.900 --> 01:56:31.940 are released in the cell and secretory vesicles which fuse the last membrane. Now if you knock out 01:56:31.940 --> 01:56:39.780 E-protein expression this pathway is still viable but you reduce virus yields by about two logs 01:56:39.780 --> 01:56:44.900 and if you knock out the M glycoprotein expression or M and E together you don't make any virus 01:56:45.460 --> 01:56:51.620 so M protein will stop without the M protein nothing happens the E-protein will still 01:56:52.660 --> 01:56:57.860 get some viral production I wonder what they mean by that I guess it's just detectable RNA in the 01:56:57.860 --> 01:57:04.100 supernatant. So our approach to de-optimize SARS 01:57:04.100 --> 01:57:14.740 coronavirus the SARS coronavirus genome and to attenuate pathogenesis focused primarily on the 01:57:14.740 --> 01:57:21.460 E and the M glycoprotein genes of about 350 amino acids in total. The strategy we used was to 01:57:21.460 --> 01:57:26.900 progressively increase the number of de-optimized codons so we started with serine to produce a 01:57:26.900 --> 01:57:34.180 D-serve virus serine-loosing R gene to produce the SLR de-optimized strain or a five-set DSLR 01:57:34.180 --> 01:57:39.700 VA strain basically the idea is you create a real stat where you're increasing the number of 01:57:39.700 --> 01:57:44.020 de-optimized residues and turning down expression of critical proteins needed for release. 01:57:45.220 --> 01:57:50.100 So that's an interesting strategy right he's changing the codons 01:57:50.420 --> 01:58:00.180 not changing the codons like to different different amino acids but he's changing the amino acid 01:58:00.180 --> 01:58:08.500 codons so that they're not as they're not what they were in the virus which he assumes is optimized 01:58:09.460 --> 01:58:16.580 which I think is a pretty cool thing because the the assumption for the mRNA 01:58:17.220 --> 01:58:27.780 transfection is that the viral codon selection the the actual codons used by the virus are not 01:58:27.780 --> 01:58:33.380 important and in fact we can change them to whatever we want so that we get more protein 01:58:34.180 --> 01:58:41.300 and we can even chemically alter the RNA with M1 pseudo-uridine alteration so that's fine 01:58:41.300 --> 01:58:49.460 we don't care about that either even though here Ralph Barrick is saying that just by changing 01:58:49.460 --> 01:58:58.260 these synonymous codons for one or two or five amino acids he can progressively attenuate 01:58:59.460 --> 01:59:05.940 viral production he can hamper viral production he can handicap the virus just by changing its 01:59:05.940 --> 01:59:13.620 codons this is pretty antithetical to the idea that Moderna can just change those codons and 01:59:13.620 --> 01:59:20.980 it doesn't matter at all right that's that's pretty impressive this is just a cartoon to show 01:59:20.980 --> 01:59:26.340 amino acid sequence and the wild type virus sequence here so like in the three set at the 01:59:26.340 --> 01:59:32.180 searing residue which was optimal in the case of SARS we just change it to the most 01:59:32.420 --> 01:59:43.460 rare codon if you look at the statistics here in general in the wild type E and M gene they're 01:59:43.460 --> 01:59:50.420 about 39 codons that are de-optimized in the one set this increased to 52 the three set 94 and the 01:59:50.420 --> 01:59:58.900 five set 134 so they're only de-optimizing the codons of the M or the E protein which is even more of a 01:59:58.900 --> 02:00:06.180 subtle modification think about that so look at the ct ctb values that Eckerd just talked about 02:00:07.140 --> 02:00:09.780 by these values here so these are very minimally 02:00:13.140 --> 02:00:20.820 on the negative side of the codon pair usage we also made random controls where we scrambled 02:00:20.820 --> 02:00:25.780 the codon usage much like Eckerd did retaining the wild type genome organization lightless and we 02:00:25.780 --> 02:00:33.300 made three set in the five set this is we made these in a mouse adapted background so they would 02:00:33.300 --> 02:00:40.660 be pathogenic in mice mouse adapted growth is shown here black the blue lines show the searing 02:00:40.660 --> 02:00:47.860 de-optimized viruses which also reach high titers by about 24 hours post-infection if you look at 02:00:47.860 --> 02:00:52.020 the three set mutants however you see about a log log and a half reduction as compared to wild 02:00:52.020 --> 02:00:57.940 type the three set randomized virus and the five set randomized virus in this case i'm showing 02:00:57.940 --> 02:01:03.060 two different plaques and the five set randomized viruses through just about like wild type so just 02:01:03.060 --> 02:01:07.940 like Eckerd had reported if you randomized the sequence it really has very little impact on 02:01:07.940 --> 02:01:15.460 replication but the optimization does affect final yield now in contrast to what Eckerd talked 02:01:15.460 --> 02:01:22.100 about we actually deoptimized in the middle of the downstream ores which would potentially affect 02:01:23.140 --> 02:01:28.340 critical sequences that would affect RNA synthesis we're very careful not to knock out any known 02:01:29.140 --> 02:01:35.780 sequences regulatory sequences that make messenger RNA however both the searing and the SLR mu knocked 02:01:35.780 --> 02:01:40.740 out messenger RNA six expression which you can see right here these actually were the messages 02:01:40.740 --> 02:01:46.340 the genes the messages that encode the gene so this is supposed to be a northern blot where 02:01:46.340 --> 02:01:52.820 the darkness is supposed to show you RNA so this isn't a very clean blot there's an awful lot of 02:01:54.180 --> 02:02:01.540 what is this i mean holy balls you're not getting a clear signal here certainly i mean 02:02:02.100 --> 02:02:12.020 i don't know what i see here but that's not what i would expect to see um he's looking for sub-genomic 02:02:12.020 --> 02:02:18.660 RNAs i guess so this would be the full genome up here i don't know that that's what i would assume 02:02:18.660 --> 02:02:24.580 um it's two hours and i don't think he's going to say very much i'm just going to leave it go but i'm 02:02:24.580 --> 02:02:28.980 going to say very much i'm just going to leave it go but i might put it on a little faster and i 02:02:28.980 --> 02:02:36.420 got to get to bed holy cow it's my birthday today tomorrow so we deoptimized so we actually found 02:02:36.420 --> 02:02:40.020 some evidence of long-range RNA RNA interactions that are regulatory elements that we're trying to 02:02:40.020 --> 02:02:45.620 decipher the five set the optimized trains showed no cpe in culture if you develop primaries to the 02:02:45.620 --> 02:02:49.940 five prime end of the genome and the leader sequence and a primer down here in the message six 02:02:49.940 --> 02:02:53.460 that encodes work six you can actually identify leader containing transcripts which are signatures 02:02:53.540 --> 02:02:57.060 of the messenger RNAs that are made during infected cells in these cultures you can see evidence of 02:02:57.060 --> 02:03:02.260 message seven i'm sorry message six five four and three this is a day one post-transfection 02:03:02.260 --> 02:03:06.020 even by day ten you can still see these transcripts infected cells and they continue at that level 02:03:06.020 --> 02:03:09.540 with about five passages at five-day intervals but you actually never can actually develop 02:03:09.540 --> 02:03:13.140 a virus you can never plaque a virus you can never actually show a virus is there producing cpe 02:03:13.140 --> 02:03:19.140 so that's interesting so now he's looking at how the RNA signal develops over days of passage 02:03:19.940 --> 02:03:24.580 and he he shows how the robustness disappears i think that's kind of cool 02:03:25.460 --> 02:03:32.500 um i don't see a control here though i don't see how the RNA sustains over in days when it's 02:03:32.500 --> 02:03:39.620 not attenuated that would be a nice control to have here right um anyway i think this is where 02:03:39.620 --> 02:03:45.060 he's going to end it he's probably just going to close he had deoptimization he's going to show 02:03:45.060 --> 02:03:50.980 these three different ways that they did it and the sites that they changed and then he shows 02:03:50.980 --> 02:03:58.260 pathogenesis of these things in the mice he made a mouse adapted version of it for aged mice um 02:03:58.260 --> 02:04:03.220 it's an it's an it's an oh these are all papers that we've looked at before they're all relevant for 02:04:03.860 --> 02:04:12.900 the clone discussion because the clone discussion puts us in um the context of the pandemic and 02:04:12.980 --> 02:04:19.300 trying to explain what's happening here i want to just leave you with the slide that i put up 02:04:19.300 --> 02:04:30.260 earlier um with regard to alina chan um and kevin mccurnan's objection that uh maybe there should be 02:04:30.260 --> 02:04:37.940 some some so ask yourself why he's making six-hour scooby-doo videos instead of downloading data 02:04:38.420 --> 02:04:43.700 from ncbi and find a signal for his hypothesis in real data if the clones offer some advantage 02:04:43.700 --> 02:04:51.620 for dr evil surely there is evidence of reduced mutation rates to prove this and i would humbly 02:04:51.620 --> 02:04:59.140 submit that actually alina chan and employee of the broad institute for whom your uh old boss is 02:04:59.140 --> 02:05:04.980 now the director for quite some time um has data from the beginning of the pandemic that seems to 02:05:04.980 --> 02:05:15.300 show a much lower mutation rate in the sars 2 sequences than those found during a similar time 02:05:15.300 --> 02:05:22.980 frame in the sars pandemic which might be the reduced mutation rate you were looking for um 02:05:22.980 --> 02:05:28.340 we have a long ways to go we got a lot of stuff to talk about but i'm not really worried the reason 02:05:28.340 --> 02:05:32.500 why i'm not worried is because people have lose in their minds and when people lose their minds 02:05:32.580 --> 02:05:37.380 it usually means that you're right over the target have never seen so many people lose their 02:05:37.380 --> 02:05:42.900 mind at once and i've never seen so many people do it that weren't doing it together before 02:05:43.700 --> 02:05:49.460 um people are coming together and retweeting and talking to one another and patting each other 02:05:49.460 --> 02:05:55.700 on the back that heretofore have never really interacted in that way before and that's pretty 02:05:56.020 --> 02:06:03.780 special not only that but it's brought together a few of these dream team members and uh i think 02:06:04.340 --> 02:06:14.500 the spontaneous sort of confluence of efforts are going to become obvious what everybody's 02:06:14.500 --> 02:06:19.700 arguing about is we all got to come together because because because i'm making exactly the 02:06:19.700 --> 02:06:24.740 opposite argument i don't need to come together with anybody because if we are fighting for the 02:06:24.740 --> 02:06:31.140 same things then our our efforts will buy necessity and buy natural 02:06:32.660 --> 02:06:38.100 by natural order where they will they will become a confluence of effort toward the same goals 02:06:38.740 --> 02:06:45.540 my goal is to have our children realize that the vaccine schedule in america is a criminal 02:06:45.540 --> 02:06:52.420 enterprise my goal is to have our children realize that virology especially RNA virology 02:06:52.420 --> 02:06:58.180 is wholly dependent on cloning technology and in fact a pandemic could not have occurred in 02:06:58.180 --> 02:07:02.820 the way that they said it is no matter how much they insist that the sequences are here and not 02:07:02.820 --> 02:07:09.220 there or that they're real or they're not they are lying about the fidelity and that's all that i 02:07:09.220 --> 02:07:15.940 can say and and all this this insistence is not good enough they need to start teaching because 02:07:15.940 --> 02:07:21.780 that's what we're doing that's what we've been doing for three and a half years no one else is doing it 02:07:21.780 --> 02:07:26.100 just us just here and so if you like what we're doing you like what we've done 02:07:27.060 --> 02:07:32.180 please support our work go to get your own biological and subscribe share the work if you can 02:07:33.700 --> 02:07:41.940 we are gaining momentum tomorrow is my birthday but i will be on i'm going to try and be on as 02:07:41.940 --> 02:07:46.500 often as literally as often as possible i don't know if it's going to be every day or not 02:07:47.380 --> 02:07:53.940 um but i really am targeting every day but i also have other things that i'm trying to get 02:07:53.940 --> 02:07:59.060 regular with including the sub stack i don't think i'm going to make a sub stack and transcript of 02:07:59.060 --> 02:08:03.540 this one but i want to do it for a lot of them and so i don't know if you've noticed it or not but 02:08:03.540 --> 02:08:11.140 there is a sub stack now the sub stack has videos that are on vimeo um those videos i've added 02:08:11.700 --> 02:08:19.780 uh i've added subtitles to and so that makes them also kind of um better to watch you can also 02:08:19.780 --> 02:08:23.860 share them a little bit better the sub stack maybe is a better way to share them because people can 02:08:23.860 --> 02:08:32.340 scan the the the transcript as well and so wolf gang wodocks interview is up there um my presentation 02:08:32.340 --> 02:08:38.660 to the uk doctors is up there um and uh i'm just going to keep trying to do it every time i get a 02:08:38.660 --> 02:08:43.860 stream that i think is good enough or is worthy of it um we're going to do a sub stack on it i 02:08:43.860 --> 02:08:49.460 don't think i'm going to do this one because tomorrow i'm going to try and do if all goes well 02:08:49.460 --> 02:08:55.460 i'm going to try and do a pretty decent show tomorrow um but it's a birthday and i've got two 02:08:55.460 --> 02:08:59.460 basketball games so i don't know there's going to be a show and it's don't know if it's going to be 02:09:00.660 --> 02:09:06.100 slam-bam crazy show or if it's just going to be an average show like this one was but anyway 02:09:06.820 --> 02:09:12.180 we've got the momentum going again or at least i got it going tonight and so hopefully i can get 02:09:12.180 --> 02:09:19.540 some sleep and uh get it going tomorrow again and uh we'll get this this pace up to speed thank you 02:09:19.540 --> 02:09:35.460 very much for joining me and i will definitely see you again tomorrow 02:09:49.540 --> 02:10:16.020 i have no responsibility for the current pandemic