Thank you, Roya. And thank you, Roya, Christian, and Rudy for organizing this really wonderful meeting and for including me in it. I'm going to be talking about joint work with Chuck Nobler, who had hoped very much to be at the meeting. And I'm going to be talking about the work of several former students and current students. Avi already mentioned Jago Pal. I'll be showing some of the work of Ruben Kedena-Nava, who's professor now in Mexico. Mauricio Comasco is here. Alan Rhine is lab at the NIH now. Reese Garmin is here at Harvard, and Vinayana Harnes' lab. Christian Byrne is here. Current graduate student, Adesia Zizgalshani is a PhD student and postdoc in the group. I have a little square that pops up on my monitor. One out of three times, I turn on my computer, and it'll be there throughout the talk. Sorry about it. I'm glad it's not a bigger one. So Chuck and I have appreciated from the beginning that we've had the great benefit of a brave and talented group of graduate students who have to learn molecular biology and biochemistry on their own and who are willing to try out our ideas about the several viruses we're working on. Avi already mentioned that we've been doing joint theoretical work almost 40 years now. And he mentioned these particular theory students. So one of the ways we started trying to understand viruses from the beginning was to contrast RNA viruses with DNA viruses. When I talk about RNA, I will mean single-stranded RNA even though I won't say it explicitly. Similarly, when I talk about DNA, it's double-stranded DNA. So the question we found helpful to ask was to understand the difference between life cycles and many features of RNA versus DNA viruses. Wouldn't it be useful to think about the differences between RNA and DNA, the genomes themselves, as physical objects? And we've already heard a lot about how DNA, double-stranded DNA, is strongly confined in viruses. It requires a lot of work to package the genome. The release or delivery of the genome is spontaneous. RNA viruses, the packaging of the genome, the formation of the virus particle is spontaneous. You're going to a lower free energy state. Suggests you'd have to do work to get the genome out. We want to understand these things in terms of the extent to which an RNA gene is a different physical object from a DNA gene. I'll show you some examples of how explicitly I mean that. I want to emphasize that we find it useful to talk about the genotype and the phenotype in the same breath, namely the RNA genome corresponding as it does to a certain sequence of nucleotides and therefore coding for certain proteins. Clearly that molecule is the genotype, but its size or particular secondary structure features that bind proteins in certain ways, that's the phenotype of the virus.  That's the usual phenotypic characterization. In this case, you're asking, what does the genome look like as a physical object? So that's the phenotype. We've seen this picture several times. I show it because I want to contrast it with the situation you have for RNA viruses. Here the whole point is that DNA is strongly confined by being packaged in its protective capsid. The size of the capsid is small compared to the size of the genome. The DNA contour length is orders of magnitude larger. Significantly, the persistence length of DNA is smaller, of course, than the contour length. That's why this behaves on a larger length scale like a semi-flexible polymer. But the persistence length is larger than the radius of the capsid, which means the DNA is strongly bent all along its length in addition to being strongly crowded on itself. And that's what gives rise to the high forces, pressures, and the large amount of work you have to do to package the DNA. In the case of RNA viruses, again, you've seen several introductions to this. Avi just mentioned, and I think in previous talks, this example was given. There's something special about 3,000 nucleotides for this virus, namely, that's the typical gene content or nucleotide number associated with each one of the identical capsids that protects different parts of the genome. This virus could have evolved to have all its genes in one molecule in a bigger capsid, but there are many important reasons why the plant viruses need to be small in at least one dimension. And so you commonly have multiportide genomes and protect pieces of the genome one at a time. We've just heard about what allows this packaging to occur spontaneously, namely that the RNA, unlike DNA, is already compactified before you add capsid protein and try to protect it. This is a cryoelectron micrograph from a Jacob Paul's work. To make the RNA molecules that we package, we make them by in vitro transcription from the cDNA. And so when we want to work with the viral RNA molecule of CCMV that's a little over 2,000 nucleotides long, we transcribe it from a piece of DNA that length. And so conveniently, you can ask, how differently do these two molecules look? They have exactly the same genetic information, but one is in DNA form. This is the 2117 base pair long template from which we make the 2117 nucleotide long single-stranded RNA. And you see the RNA molecules are branched. They're compactified by the branching. And you have this amount of genetic information in very efficient form. DNA is a less efficient way from a space point of view to store genetic information. DNA, double-stranded DNA, has many advantages, obviously, chemically and otherwise over single-stranded RNA. But as physical objects, we see the difference in the phenotype of this genetic information. Here is a picture of one of the three molecules of the virus, CCMV, RNA 2. It's almost 3,000 nucleotides long. We have a purified sample of it. We take pictures of it and try to sort out branching characteristics and so on. But we can put in a scale bar, determined in the usual way from the microscope. This is the best scale bar of all. We throw in a little bit of the purified virus. We know that that's 28 nanometers. And you see that indeed, as you've seen in previous slides, the RNA that is packaged by protected by the capsid protein you add to a solution of the purified RNA under assembly conditions, it's already about the right size. In contrast to the situation for DNA, I want to speak briefly about the two-step in-vitro self-assembly protocol that was worked out by Ruben Briceau in ultimately clarifying it as follows. We mix the purified capsid protein and the RNA of interest. It can be viral RNA or non-viral RNA, ranging from hundreds to up to 10,000 nucleotides. Mix them in physiological buffer and then dialyze against lower pH. What happens at neutral pH is that the RNA is saturated by capsid protein, and you lower the pH heading towards the isoelectric point of the protein, and you enhance the lateral interactions between the proteins that are bound on the RNA. And then you have the formation of capsids. And you analyze the assembly products with gel, sucrose gradients, or looking directly at what you have. These are pictures from Brice's work where he wanted to check that indeed you have to lower the pH to at least six from neutral pH before the protein-bound RNA complexes form proper variants or virus-like particles. Happens in negative stain, lectin microscopy. By adding urinal acetate, you're lowering the pH by several units. So you can take the neutral pH assembly mix where we know from cryoAM that you don't have properly formed capsids. You just have RNA saturated by protein. When you prepare your EM sample, you are actually lowering the pH. And that's why you see capsids. What we learned, again, through the work of Ruben Cadena, Mauricio Comas-Carzi, and Ries Garment, is that if you have a sufficient excess of capsid a super stoichiometric one. So for 3,000 nucleotide long molecule, instead of needing only 180 proteins per RNA molecule to give you a T equals 3 capsid containing that molecule, to get complete packaging of the RNA, you need more like 300. So corresponds to a 6 to 1 mass ratio of protein to RNA. And we call it the magic ratio because it allows complete packaging no matter what the RNA is if you have that mass ratio. And it happens to correspond, not coincidentally, to the mass ratio that allows the cationic charge on the n-termini of the proteins to be equal to the anionic charge on the RNA. And if the RNA isn't too short or too long, you get complete packaging into a wild type T equals 3 capsid. If it's shorter than that, you get smaller capsids. If it's much smaller than that, you get many copies of RNA per capsid. If it's longer than 4,000 nucleotides, you get the wild type structure, the T equals 3, but you get two or more capsids sharing a single RNA or multiple. Learning how much capsid protein you need to completely package an RNA allowed ratio to do well-defined competition experiments where you take equal masses of two RNA molecules whose relative packaging efficiencies you would like to determine, you take equal masses of them and add six times as much mass of protein as you have mass of either one of the RNAs. So now you have a 3 to 1 mass ratio of protein to total RNA, it's enough to completely package one of them, but not both of them. So now you have proper grounds for a competition and you go through the same two-step packaging protocol. Perhaps I didn't mention before, you add RNAs to get rid of any unpackaged RNA. And then you analyze your assembly products and see what fraction of the virus-like particles of one RNA and what fraction of the other. Through a lot of work of that kind, we learned that indeed, viral sequences are packaged more efficiently than equal-length non-viral sequences. And we interpret that, understand that in terms of the viral sequences giving a more compact structure, which can be more easily accommodated by the protein that prefers a certain curvature and T equals T structure. We've heard from Raiden about the importance of packaging signals. Obviously, the viral sequences have them, non-viral sequences don't. So after sorting things out along these lines, we thought we should go outside our comfort zone and ask, what happens when you take an unnatural RNA, like a single-letter RNA, that has no possibility of forming secondary structure? It has no self-complementarity. We picked Polly U because of out of all the single-letter RNAs, it's the one that has the weakest base stacking so you don't even get any helical ordering of the single-stranded polymer. We guessed that for all the reasons I've been talking about, it would not be an efficiently packaged molecule and certainly couldn't compete with normal composition RNA, little viral RNA. And it turned out to be a very interesting experiment because we got nothing of what we were expecting and we're still trying to understand it. It's hard to prepare monodisperse samples of Polly U. See, it is synthesized, you have a broad distribution of sizes, you can fractionate your sample into very roughly monodisperse plus or minus 50% samples. So instead of talking about 2,000 nucleotide long Polly U, we talk about Polly U in this range, this range, this range, and so on. And we take each of those samples. This is the work of Christian Baron. He doesn't have a poster on it. He has a poster on 3D reconstructions of CCMV and BMV RNA and capsids, actually just BMV for the reconstructions. We take each one of those samples, carry out the in vitro packaging, and find our first surprise that we don't see any wild type T equals 3 capsids. We only see T equals 2 sized capsids. Bogdan, looking at you, I remember years ago, when we mentioned T equals 2 structures, we saw, I'll come to them in a moment, when we're looking at short RNA molecules with normal composition going up to longer ones, generating doublets, triplets, quadruplets, and so on, showing that when RNA is shorter than 2,500 nucleotides, you get smaller than T equals 3 structures. We call them T equals 2. You said you don't know that until you do a 3D reconstruction. We know from Christian trying to do a 3D reconstruction that we have T equals 2 sized particles, but they're not T equals 2. They're trying very hard to be, but they don't manage. That's somewhat polymorphic, a little anisotropic. But again, every one of these colors is a different size of polyu, and every one of them gives a distribution centered around T equals 2 size in stark contrast to the wild type virus, or we would get the same size distribution for a normal length, say 3,000 nucleotide, normal composition RNA. We again see a succession of multiplets, and they set in at the same lengths as for normal composition RNA, but you never see bigger capsids than the T equals 2 size capsids. Then Christian did competition experiments between polyu and viral RNA, DMV, RNA1, which happens to be the most efficiently packaged molecule when you use CCMV capsid protein. It's more efficient than CCMV RNA1. But it loses in a competition against polyu. Here it's easy to analyze the assembly products. You don't have to carefully put different fluorescent labels on the RNA molecules because you're getting different sized particles, and you just count the relative numbers of the different sized particles, and you find that there are more T equals 2 size particles than T equals 3. And you confirm with the usual labeling experiments. So not only do you have the surprise that you get a non-wild type virus like particle, not at T equals 3, but it turns out that polyu is more efficiently packaged than viral RNA. Is it really packaged or it's a cluster? It is packaged in terms of what it looks like in an electron micrograph, and it's protecting the polyu against RNAs. Those are the criteria for packaging into a virus like particle. So it raises the question, what determines the size of capsid that will be formed when you take a certain capsid protein and add RNA of different lengths and sequences to it? And now, of course, we're taking a broad range of lengths. We're also taking a dramatically broad range of sequences, including polyu. So I'm going to be contrasting a single-letter RNA, like polyu, where you have no possibility of sucking the structural formation, with what we call normal composition RNA, whether it's viral sequence or not, you have comparable numbers of each of the four letters. It's easy to answer this question, what determines the curvature. You just think of all things that might determine the curvature, and you start listing them. It's the intrinsic or spontaneous curvature of the capsid protein. So it's the nature of the protein interactions. But we also know that it depends on how the conformation of the protein is affected by interaction with RNA. So here's another factor. We also know it's determined by the size of RNA, because when we go from short RNA to long RNA, we go from T equals 1 to T equals 2 to T equals 3 type structures, bigger and bigger capsids. But I remind you, we get different scenarios for different kinds of RNA and different kinds of anionic polymers. For normal composition RNA, you go through this progression of sizes, and then to multiplets involving the biggest size. The first work we did was with polystyrene sulfonate before we started packaging RNA with CCMV capsid protein. And again, we found there a progression from smaller than T equals 3, T equals 2 sized VLPs to T equals 3. With polyu, we find a very different result. You never get to the wild type T equals 3 structure, but you do go through the progression from singlets to multiplets. Kristin is presently looking at poly A and sees something similar to poly U, but we have to look more carefully at it. So here is where I want to go back to normal composition RNA and use what we've learned about packaging of RNA by capsid protein that's capable of in vitro self-assembly from purified components, namely BMV or CCMV capsid protein. I want to go back to what we've learned and ask how can we use it to develop the best gene delivery system. And so I want to go over some of the requirements for a good gene delivery system and how the answer to the question, what's the best system, will come from looking at the differences between RNA and DNA genes. RNA genes, in contrast to DNA, can be packaged spontaneously if we use a promiscuous indiscriminate capsid protein like CCMV or BMV, which doesn't care about whether it's packaging viral RNA or heterologous RNA or even synthetic anionic polymer. Now we get into a little biology. RNA genes, most of the RNA viruses that have been mentioned so far, get to HIV soon, and that's very different. They're not just RNA viruses, similar standard RNA viruses. They are positive sense RNA genomes, namely there are messenger RNA molecules that are directly translated by ribosomes in the cytoplasm of the infected cell. So to a significant extent, the nucleus of the infected cell is never involved. The viral life cycle is much shorter because as soon as the genome is made available to the ribosomal machinery, you have gene expression and viral replication. And then another thing that's special about RNA is that it can be self-replicating. And I'll explain what I mean by that and how we take advantage of it with the DNA virus. DNA gets into the nucleus. You have transcription of the viral genes. You have the needed viral gene products like capsid protein and so on. How do you replicate the genome? You use the DNA polymerase of the host. DNA replication is not an exotic thing for the host cell. But RNA replication is. So genomes of positive sense RNA viruses code for an enzyme that wouldn't be found in the cell otherwise, namely an RNA-dependent RNA polymerase. It's an RNA replicase. And that replicase messenger RNA codes for the enzyme that replicates it. The whole genome is replicated by the RNA-dependent RNA polymerase gene product. That's one of the few genes of the RNA virus. So here's our strategy. You recognize this genome? Arguably, the simplest genome organization of an virus is this one, where you have two open reading frames. Alpha viruses have this structure. One open reading frame for all the nonstructural genes. So most importantly, those genes include the genes for RNA replication. And then one open reading frame for the structural genes. It would be capsid protein for a naked virus and capsid protein plus membrane proteins for an envelope virus like the alpha viruses. What's special here is that this genome is directly translated. There's a stop code on here so that the only gene products that are directly synthesized are the enzymes that will make up to a million copies of the genome. It's a very powerful strategy for a virus to make a million copies of itself before anything else happens. Significantly, to get structural protein gene products, you need to have RNA replication. So when you make minus strands of the full length genome, from those minus strands, you make full length plus strands. Those are the copies of the genome you need for the next generation of virus. But you also start transcribing the minus strand of the full genome at this junction here. And you make millions of copies of the messenger RNA for the structural proteins, of which you need a huge number. So I argue it's the most powerful way to get brought to evolution by these viruses to get high levels of expression of protein. And that's what we want to take advantage of. Basically, we want to, an RNA viral genome like this, cut out the structural genes, replace them by genes of interest, which could be, at first, reported genes to quantify what's happening, and then viral vaccines, cancer vaccines, anything you like. We're basically inserting into the viral replicon a gene of interest in taking out the structural genes of the virus so that you don't have an infectious reagent. We need to find an RNA virus whose replicase genes are short enough to be packaged by CC and B-capsid protein. That's our next requirement. And we found an insect virus, notomura, very closely related to flockhouse virus, whose genome has a related but different structure to what I just showed you. Namely, it's a two molecule genome. And here's the open reading frame for the replicase genes and here for the capsid protein, molecule 2. So the two molecules are co-packaged. And you make the replicase, which makes a large number of copies of both molecules. And this is the replicon. It's only 3,000 nucleotides, which gives us 1,000 to 1,500 nucleotides to work with for inserting genes of interest. And that's just what we do. So the RNA is dependent RNA polymerase, which is protein A in the case of notomura. We leave intact. We add a gene of interest. We add sequence for self-cleaving peptide because we want the RDRP and the gene of interest to be functional. We make them as they are made as a polyprotein and then cleaved. And then these are pictures and analyses by Rhys Garmin. With the first construct we made of this kind, while it was still at UCLA, where the reporter gene is an EYFP, the total length of the viral replicon plus inserted gene is about 4,000 nucleotides. And you get nice RNAs resistant, thermally stable, aggregation stable, T equals 3 VLPs. Remember, we've got the replicase gene from an insect virus. We're packaging it in the capsid from a plant virus. And we want everything to work in mammalian cells. One of the reasons we picked notomura was not just that it had the right replicon length, but it had been shown to be active in mammalian cells. And we checked that. This is work out of C. Aziz-Kolshani. By transfecting these in vitro reconstituted particles with the fluorescent protein reporter gene into mammalian cells and seeing high levels of fluorescence. We've checked it since in many ways in many different cell lines because we want to use this as a gene delivery system. Five minutes? One minute. OK. Thank you. Just checking, not only that you get a high level. In this case, it's a Luciferase reporter gene. Because of the RNA amplification, you get expression for many days. This is the messenger RNA form of Luciferase. This is the replicon form of Luciferase. So not only are you amplifying the messenger RNA for Luciferase, you're doing it over the course of many days. And that was work of Adam Biddlecombe. So this is, in fact, my last slide. And I want to summarize what we've learned and where we are. We choose genes in RNA form because we can in vitro package them, which means we can protect our genes that we want to deliver, taking advantage of the flexibility and compactness of RNA genes versus DNA genes. We use the fact that certain plant viruses like CCMV have risk in discriminant capsid proteins that will completely efficiently package the RNA replicon gene. We use the self-amplification strategy of positive sense RNA viruses and take advantage of the fact that those particular replicases work in mammalian cells. So we haven't created an artificial virus, but if you like a hybrid virus, for the reasons I talked about, I'll stop now and I hope there are questions and we can discuss these things further. Thank you very much.


Hey, Bolo Buddies. Thanks for watching. Okay. If you are new here, I am Courtney and I talked to you about Bolo items items You can buy low and sell high these items are a part of my 50 unexpected Bolo series and basically what I do is I share Bolo items in my Bolo Buddies Facebook group and after Usually probably I don't know two to three weeks after the items in I know a lot of times people will like Look at the listings and then forget to go back to see what they finally sold for so I'm making a video That's gonna share with you those items after the auctions have ended So these are big money Bolo items a lot of them are unexpected but if you're in my Facebook group you may have seen them before but this is a video refresher and Usually in Facebook groups like you don't see every post anyway. You just get they're hit and miss what you see So this is part four if you haven't seen part one two and three definitely go check those out And we're gonna get started with these big money Bolo's and I do mean big money So these are items that you definitely need to be looking for when you're sourcing I know that I am and I started the Facebook group to learn and I Started the YouTube channel to learn because I used to only sell clothing So I am now a Bolo finder. I love finding Bolo items, but I sell hard goods now So let's get started The first item is an Item that I'm not gonna share with you And the reason why is is because when I went back to look at the item it said Relisted this item which means The buyer probably didn't pay and it probably didn't sell for that much. So this item is Relisted It did sell for 560 but I'm guessing it was non-payment So if you click on Relisted item you can see here that the item ended up selling for $455 With 28 bids So here's another thing you can do just a little teaching moment We're gonna go into their feedback and we are gonna type in Romeo and see if feedbacks been left So no feedback has been Left yet, but the sale just ended on March 7th So it probably hasn't been delivered yet So that's not a good way to check But if you're looking at another item and you want to see if the sale went through That's a good way to go check because eBay now allows us to search feedback with a keyword, which I love All right, so let's get into these 50 Bolo's the first item are these vintage fine quality purple lavender jadeite Clip on earrings and they put clip earrings Yeah Clip they're calling them clip. They almost look like those don't look like clip on earrings. Do they I? Want to see what they look like open? They look like shoe clips or something Anyway, they're selling them as clip earrings for $356.51 22 bids buyer paid shipping The next item is this blithe doll and I think I've shared this one before but I asked and Somebody said there was something on the back that you pull To make the eyes change colors, but I'm not seeing that on her back Is it lower? Oh, oh This one's on her head So does it vary by doll or maybe they meant the back of her head? But yeah, you pull on that string and her eyes change colors. Isn't that different? It's kind of cool, and it's kind of creepy all at the same time $1,586 58 bids buyer paid shipping The next item that sold is this turkey call makers past and present book and it's vintage and it does look like it's Signed so that could have increased the value of this one But I feel like they should have put like signed in the title or something like that But this sold for $555 83 bids buyer paid shipping The next item is this Lego Dragon Masters new old stock Lego set 44 bids sold for $850 and the buyer paid shipping How amazing is that for a Lego set? The next item that sold is this pyrex dish now I shared this one in my Facebook group and I had a lot of comments and People were saying that it wasn't the pattern that was special But the pattern on this shape of bowl. So whatever that means I have no idea I don't even know if they were correct But that was some of the feedback that I got as to why this went for so much is sold for $2026 with 41 bids plus shipping. So if you guys know anything about this pyrex bowl Please put info down in the comments so that we can all go check the comments The next item that sold are these Barbies Um This actually still has 22 hours left, let me see if it updated Yeah, 22 hours left now my question for you guys is and the reason I included this is because I Didn't have 50 so I had to pull some that were active and I was hoping they would end by the time I recorded But they didn't Would you guys have listed these individually or as a lot like this? I feel like I would have listed them individually But I'm looking at what the buyer is getting or I mean the seller is getting and it's at 865 dollars So, I mean two four six eight ten twelve dolls So, I mean they're still getting a pretty penny. So I don't know I'm not a Barbie I don't know tons about Barbies I like selling Barbies, but I'm still learning. So like I know that's a bubble cut up there and But I mean that one's got green ear and if you guys want to learn more about like how to Like clean the Barbies up and different things like that about Barbies go to my reseller treasure. Is that it? I Always mess up her name Think that's it my reseller treasure Let me check Instagram. I've got that pulled up on my phone I can find her real quick. Her name is Brandy and she is fantastic, but I can never Remember or I always question myself. Yeah, my reseller treasure. I don't know why I was questioning myself That's her YouTube channel. Definitely check her out The next item is this oh It ended But they didn't pay so $223.50 it was one shoe only and It sold with 38 bids. So they re-listed this item. Let's go in and look and It's currently at $40, but it is one shoe. Apparently it's some sample shoe But I was like why would they use the tag as the main photo and somebody in the group said because that tag Has all the important information So, I don't know. I guess people can buy one shoe. I've sold a baby shoe that was one shoe But it was like an antique Like Victorian style boot Hard to find somebody may have had the match The next item that sold is this vintage and this one still has 21 hours also 1966 peanuts Vinyl hunger for pilot. It's a little Snoopy figure. It's currently at $110 and the buyer will pay the shipping on this Again, I don't typically show you the active auctions. I wait until they end for this video, but I was a few short So that's why I did it that way The next item that sold is this Vintage Bonn Bell roll-on Shiner. It's got 16 hours left. It's currently at $80 Do any of you guys remember using this as a kid? And was it $88 great? I just want to know The next item that sold is this rare 2010 vintage So that's what's up for discussion in my Facebook group. They're like that's not vintage and I'm like, yeah 2010's really not vintage retired is a better word. They used retired here But I don't know what this is it says yarn doll But it's not yarn. It's like some sort of figure. I don't know if it's resin or Wood or what it's made of But it is a big money item There's actually another one that I saw that was listed and it is also going for a lot of money So people must collect them It's currently at $296 with 33 bids and the buyer will pay the shipping The next item was sent to me from a person in my Bolo buddies Facebook group so I invite you guys if you ever have a eBay auction with bids that you picked up cheap and Are selling for a good profit? Please feel free to message me on Facebook Using messenger with that link and I will share it for you guys and get you some exposure for your item and it also helps people in the Facebook group learn and as if I continue this series which I hope to do I Plan to do unless somebody is like I hate it or multiple people hate it But so far the I'm getting really good views on it and I'm getting good feedback. So I think I'll continue it But somebody sent me this in the Facebook group. They got it for $5 at a yard sale. It is a Homer Laughlin Fiesta small vase and it sold for $69 plus shipping and that's what it looks like on the bottom So definitely check out fiesta if you see it just look it up and see what they're selling for The next item is this 1960 vintage Rolex hang tags It's like a wax seal hang tag. You guys this sold for $829 58 bids buyer paid shipping now. I Know with the Rolex Boxes people have told me that you can't sell a Rolex like original case. I don't know if that's true I've never researched it. I don't know if this is considered the same thing So my advice to you would be call eBay and ask them before you list it or Don't put it on eBay sell it somewhere else because you definitely don't want to compromise your eBay account And again, if I ever show a Bolo item There's always a chance that somebody got away with something that they weren't supposed to because there's so many items on that Vero list So definitely do your homework. I Try not to list things that I know are Not allowed on eBay and if I know they're not I will tell you guys like if somebody shares it on my Bolo live Show I'll be like, oh, I wouldn't list that. I've heard some things But again, it's just it's kind of one of those gray areas sometimes The next item is this lot of 222 vintage Mechanical pencils and this auction just ended Today at 326 so my screen updated there these sold for $305 so somebody had a nice collection of mechanical pencils and Decided to get rid of them. This could have been in an estate find or something like that. I don't know Things like this you can pick up pretty cheap sometimes so 30571 bids $16 to ship the next item is Another one of those lip smackers and this one has 15 hours left and it's at $222 and if you go into this seller store They have a bunch of this stuff listed. They like hit the lip smacker vintage lip gloss Whatever jackpot and if you go to my Bolo buddies Facebook group and look for this listing You can click on it and then go in and see sellers other items if you want to see what else there's a lot of Bolo's 15 watchers still on this but it's currently at $222 The next item is this vintage disneyland new Orleans Caribbean condiment jar and this sold for $453.43 bids buyer paid shipping the next item. I thought was really really cool It's a lot of six vintage 80s Reagan years white house egg roll wooden Easter so they're like wooden Easter eggs How funny is that? I mean it's not really funny, but it's kind of funny. They're from the White House I thought these would go more actually they sold for $256 and 78 cents plus shipping Here's another one where I feel like I would have sold these items individually, but again I'm not I think I've sold that Barbie head the one with the headband. I think I've sold her head only before But yeah, I feel like the buyer probably could have got more selling them individually They sold them for $446 and the buyer paid shipping The next item is this vintage 90s sailor moon disguise pin Yeah, this one sold for a lot of money I Think that this sailor moon stuff is collectible So if you see sailor moon, definitely look it up this sold for $390 plus shipping The next item. Oh, I already did this one. Maybe you're only getting 49 Bolo's. Sorry guys This I love this. I love it. So it's a uranium glass ring. Have you ever seen anything like it? It's so cool. I mean that is cool. You have got to admit that that is cool let me know if you think that the uranium ring is awesome and go into eBay and look up uranium rings. They're like a thing. I Don't know what that is CPM. Hmm. I don't know what that's testing for but this sold for $307.78 plus shipping. So if you see a green ring, I guess black light test it and see Pretty cool, right? The next item that sold is this golden astronaut moon exploration space play set Pretty cool sold for $122 and the buyer paid shipping The next item that sold is this vintage cotton batting hand quilted embroidered basket flower quilt. So it's got the flower baskets as the print I feel like my grandma had one of these for some reason but this sold for $455 plus shipping The next item is huge 520 pieces and I did look at this person's store and they do sell Barbie items So I'm really shocked that they lauded all of this up But I guess if they sell Barbies, they know what they're doing look at all those heads This is a huge lot massive just amazing Wow $1,720 is what this sold for and the buyer paid shipping The next item that sold is another vintage Barbie. This is a number two Barbie. I think She a ponytail. I can't tell I get confused. She might be a ponytail Barbie I don't know what she's called Don't quote me on that, but she does have a ponytail She's in pretty good shape Her makeup looks pretty good. She sold for $5,650 with 46 beds and Barbie collectors. Let me know what she's called because I don't know if she's a ponytail Barbie. She might not be The next item is this hard to find vintage pyrex eyes carafe with lid turquoise HTF hard to find and the reason I'm telling you that is because I didn't always know that so There may be somebody that doesn't know that so I'm just sharing that with you hard to find OAK is one of a kind this sold for $250 and 50 cents in the buyer paid shipping The next item is this vintage zippo lighter. It's a high polish chrome military. It says sold for $990 plus shipping The next item are these huge lot of viewmaster reels sold for $265 and the buyer paid shipping it looks like a lot of them have the packaging so I'm sure that helps I Don't know. I guess be on the lookout The next item that sold is this nirvana in utero t-shirt It's super rare And that's what it looks like and it sold for $3,900 plus shipping The next item is this vintage pyrex glass turquoise white balloon chip in dip set sold for $359 buyer paid shipping The next item that sold are these vintage antique Stifle junior fabric overalls preteen child never worn. So I Would say that adult clothing sells for more money So if these would have been adult size, who knows what they would have went for but they went for $427 and 52 cents plus shipping the next item is this T twist in turn TNT is twist in turn Mario flip Barbie Burnett perfect coloring 1969 no fade sold for $577 and 77 cents 47 bids buyer paid the shipping The next item that sold is this Lisa Frank vintage peek-a-boo mushroom lunchbox Like I said, Lisa Frank is a bolo if you see it. Definitely grab it. Look it up Comps should be good, but if not, you're still gonna make some money probably as long as you get it for a good price Bolo item $197 and 50 cents plus shipping The next item that sold is this rare 1986 been vintage constrictor Marvel Secret Wars action figure made in France sold for $95 plus shipping The next item that sold is this Honda service manual and it sold for $127 and 50 cents plus shipping The next item that sold are these roller skates roller skates definitely range. These are not in great condition That's what I wanted to show you and they still sold for a hundred and seventy two dollars and fifty cents You can also sell the wheels so definitely Look up skates if you see them because they can definitely be a bolo the next item that sold are these magazines and They sold for $96 57 bids buyer paid shipping these are hip-hop rap R&B music magazines And I will tell you that prime time treasure hunter on YouTube lots up magazines all the time He's done like the muscle builder like muscle building Trying to think of some of the other ones, but definitely check out his channel Check out his Instagram also prime time treasure hunter because he shares his bolo sold items there And I always see the magazines So definitely check that out this vintage B&G Bing and grotto porcelain pug dog figurine So for a hundred and forty seven fifty with fifty one bids plus shipping. There's the mark on the bottom Super cute. I would say this would appeal probably to the people that collect that brand and also people who collect pugs Vintage Disney map. That's the title sold for two hundred and thirteen dollars and fifty cents plus shipping Anything Disney look it up for sure That's cool. It's got all the different little paperwork items that go with it. I don't know if I'd Mary Wonder how Mary feels about that. All right. The next item is this 1985 vintage garbage pail kids first series Unopened three hundred and thirty four dollars forty eight bids. It's one pack guys The next item that sold is this vintage 1975 Dungeons and Dragons white box fifth printing book sold for four hundred and six dollars plus shipping The next item is this vintage Fisher price fun with food for little tykes wedding cake mixer and cookies. I Love this bolo. I love selling toys. I don't know if I would have sold these as a lot It seems like a lot to put in a box But the seller did pretty well. I don't know if they would have sold better separately. What do you guys think sold for two hundred and thirty five dollars and forty nine cents buyer paid shipping The next item is this silver tone radio model vintage as is for parts or restoration This sold for three hundred and forty one dollars and the buyer paid shipping The next item that sold is this huge poly pocket Disney magic kingdom figurines Cinderella I actually Peter Pan. I actually thought this would go for more and when I put this in my facebook group. It was much lower so it did get some um It did get more bids as it was posted Longer maybe it was like a bidding war but right before it ended But there's a this is a pretty decent size lot and there's a lot of figures And it only went for a hundred and seventeen dollars. I probably would have parted it out Um, but I don't know what do you guys think about that one? Let me know in the comments This vintage steel chainsaw trucker farmer hat sold for ninety two dollars plus shipping Trucker hats are definitely a bolo They range though from bread and butter to big money. So definitely um, look them up Vintage lace seems to be trending right now too. Um, I keep seeing a lot of those when I'm searching for auctions This one sold for two hundred and seventy five dollars plus shipping The next item is another quilt. This is the feathered stars broken star vibrant Fire brick red quilt sold for three hundred seventy three dollars Plus shipping. This is one, you know, I know I didn't know quilts went for such big money So Definitely be on the lookout. I mean I do now since I you know do Do these shows and stuff, but I feel like before All right, this lisa frank backpack An a member of my bolo buddies facebook group sent this to me and asked me to share it in the group And I did and it's an awesome bolo and they also shared it at the bolo live show, which is great. So they're definitely Excuse me getting involved and helping others learn and I definitely appreciate that This sold for a hundred and twenty seven dollars and fifty cents and the buyer paid shipping The next item that sold are these vintage nineteen thirty six alexander dolls This was another one where the I think the buyer didn't pay and it was relisted When um, it went through the second round of auction It did sell for less, but it still sold for two hundred and forty five dollars and fifty cents plus shipping And the last item is this antique shoalts and fissures sterling silver cleopatra cake saw serrated knife Great key words beautiful knife very very fancy. This sold for a hundred and sixty one dollars and thirty eight cents plus shipping All right you guys Those are the 50 Bolo's from my facebook Bolo buddies facebook group definitely go check it out if you haven't already um, again, there were a few people from um My facebook group that shared if you would like to share Definitely reach out to me on facebook if you have an auction with bids And if you miss that opportunity you can always come to the Bolo live show on wednesday night 10 p.m Eastern standard time and share your Bolo there in the chat. Um, it's a live show and its audience Audience viewer participation. So definitely come check that out every wednesday 10 p.m eastern standard time Thanks for being here. Thanks for watching. Please leave me a comment And if you know somebody who could benefit from this video, please share it out Feel feel free to tweet it pinterest it Facebook it Get it out there. Thanks for watching And if you share the video comment below I shared so I know all right you guys. Thanks so much


What's good George the boros back again with another video, so Apparently MJF Goes off in last night's media scrum. I didn't get a chance to check it out But a lot of you guys were saying I need to check it out. So that's what I'm here to do in my Luscious robe. I've been in a road all weekend. I've just been relaxing man Just been relaxing still dropping some content for you guys But you know, I just wanted to kind of relax as much as I possibly can this weekend because this week it's gonna be crazy I'm there's a good chance. I'll probably reach a hundred K subscribers this week. Hopefully so So, you know, I'm trying to set up the hundred K subscriber stream. We got the Survivor series stream. We got the Monday night raw stream probably got the Friday night stream Like we got so much stuff that we got to check out all also it's Thanksgiving this week So it's just so much going on gonna be hella busy So I knew I need to at least try to recharge my batteries as much as possible But we got to check this out man cuz MJF is on top of the world So I want to see what he said at this media scrum. Appreciate it.  I love it doesn't care. Let's talk shall we? AEW is now destination television once again The ship has been steered properly once again I love that belt by the way that AEW championship belt looks so damn good In this entire sport and it states the three letters and it dams you're an AEW I know for a motherfucking fact it ain't mox. It's m Jay That is so good and he played it up. I'm like this has to be obviously an act But he played it very very well Even though in this match and we've had this discussion, you know on twitter and on my last video MJF was pretty much still a heel Yes, but The way the match was set up the way the atmosphere was set up You would think if you never seen MJF before you would have thought he was a baby face You would have thought he was he was pretty much trying to come back from behind in in a situation to overcome Uh john moxley.  I want to earn it.  I'm a microphone in the ring. Nobody could touch me That's a fact.  Oh, yeah, and I I did see the off-air clip where he put him over He kind of put him over. He was he was really he That's one thing I could say about MJF. He was really hamming up. I'm going to earn it the legit way I'm not completely a good guy But I'm gonna earn it the legit way. I love that that that that's why I can't wait till Wednesday night I'm definitely gonna be streaming that as well, man This whole week is gonna be full of stream.  No offense grow up Let's talk a little bit, huh? Uh, MJF, what's going on with you and regal? You know, we're all really interested What's going on? How'd that link up start?  Huh with 70 I like the fact that he's selling the knee injury too I hope it's not a real thing. I hope it's not.   No, if you want to know anything about the most important man in professional wrestling You got to tune in to the mjf show.  Okay I'm gonna take a shower.  Huh, just kidding. Thank you.  Bye No It's so Oh my god Mjf man. He is he He knows what he's doing, bro. He knows what he's doing. I'm looking forward to seeing what happens on wednesday Uh, I'm glad he's the top champ now. We can really get some things going. I know I believe Ricky starts is supposed to be having a match where if he wins that match He'll be the number one contender for the aw championship. I'm not gonna lie to you I actually would be okay with that. Ricky starts is definitely someone that needs to be featured more on television You definitely go on the microphone pretty good in the ring I would love to see a ricky starts in an mjf kind of go back and forth Granted, you know, I I would probably wait on that. You know what I'm saying to build up ricky starts even more but If they were to go with that a little bit sooner, I'm okay with that I think they would have a fantastic little feud Um, and I'm just looking forward to the feuds he can have now As the champ because he's gonna do whatever it takes To keep it he's gonna do whatever it takes to keep the title Which means we're gonna get some great heel Content because there's nothing better than a heel doing everything he can to keep the title So that way when he actually finally loses it to someone it makes that person a mega star This was a great great media scrum. He just came in there talked his trash and left. I can appreciate that I'm glad he is the world champion. He is in my opinion one of their best top guys. He he has it He knows how to Really get people invested and I feel like these These mjf promos and these mjf segments and the people he's gonna feud with I think it's gonna be must-see television So I can't wait to see what they do with him moving forward. But comment down below. Let me know. Did you guys? That's the up calling me right now. But comment down below. Let me know Did you guys enjoy this little little segment of the media scrum? Let me know down below. At least this time it didn't End off in like a brawl afterwards. So that's that's a plus But uh, I appreciate all of the support you guys have shown on the channel so far Road to a honey k appreciate y'all kicking with me. See y'all next one. Peace