[MUSIC] The third scientist we are going to talk about is Elie Wollman. Elie Wollman was born and grew in the Institute Pasteur. His parents were scientists there. His parents were the people who, Who were hosts to Salvador Luria when he escaped Italy on his way to the States. His parents were deported in 43 and never came back. Elie was heavily involved in the French resistance, and, He actually knew Monod at the time. And then after the war, he went to the States to the Delbruck lab. And he was actually the first French from the Pasteur to go to the Delbruck lab. He started the move, and he also, in a way, started the move of all the Americans who went to Pasteur. His godfather was Elie Metchnikoff, hence he got his first name from his godfather. And Metchnikoff discovered phagocytosis, was recruited by Pasteur, and is a founder of cellular immunology. And then Monod was in the lab of, Monod took Wollman in his lab. And he taught Jacob how to do experiments with bacteria and phages. And they were very good friends, and they worked together. And the duo Wollman Jacob was the first successful duo in Jacob's life, the second one being the one with Monod. We're not going to discuss Jacob today. We'll discuss Jacob and Monod next week. But, He was a very interesting character. He was very much involved in the life of Pasteur. He considered himself as the last person alive who had known the Pasteurians, who had known Pasteur. He was a Vice Director of the Institute later on, and, He and Jacob had a great fun. They discovered, as we're going to see in this paper, something which they called, Coitus interruptus, which is mating between a male and a female bacteria with a precise timing of interruption. And they of course wanted to name the phenomenon coitus interruptus. But that was not acceptable by the official French academician. And they were absolutely scandalized that these young men could be so gross and crude. But that was luck. So he was in the Institute Pasteur basically from his birth. I mean to give you an idea of how the atmosphere of the Pasteur was. When Elie was born, his father Jean Wollman met Roux, who was the chairman at the time. And Roux, a first disciple of Pasteur, [FOREIGN]. So, Wollman, how is it going? Very well, Mister Director, I just had a kid and time now is pretty hard. Okay, I will give you a raise and appoint you chief of the lab, but don't mention it to anybody. I was always, things were done at the best way. And so, then things were a little bit harder. Because Wollman considered that he, Jacob, and Monod, had been the generation of the young, bright kids. Lwoff was a big daddy. Lwoff was a professor. And so when the Nobel Prize was given to Lwoff, Jacob, and Monod and not to Wollman, that strained a lot the relationship between these people who were pretty close friends. And became a little bit less friendly after that. And Wollman always felt, and his family always felt, that he'd been mistreated. On the other hand, Lwoff had studied lysogeny and had made major contribution to the field. So if you really think with a little bit of perspective, Wollman's contribution was important, but Lwoff was very important. And so it's not completely unnatural that the prize went to Lwoff, Jacob, and Monod. So today we're going to read a paper that appeared in the [FOREIGN] which is the French equivalent of PNAS in the States. Except that it's a very, Very different kind of journal. In the 50s and the 60s, a lot of papers were published in the [FOREIGN] that were then completed and published in other journals. To be published in the [FOREIGN] you had to go through a member of the Academy. And the member of the Academy had to submit it. And in this case, it was Trefouel who submitted most of the papers from the Lwoff, Monod, and Jacob, and Wollman group. And this is one of the examples, it was published in June 1955. And basically what they did here is they used a big discovery made by somebody Scottish, a guy named Hayes. Who discovered that some strains that are F+ could convert spontaneously into strains that give recombination with high frequency. Hence high frequency of recombination, HFr. The difference between an HFr and an F is at least 100,000, if not a million fold inefficiency. This is the real carrier of the genome of one bacterium that will recombine with another bacterium. Everything that had been done painfully with F, between 47 and 53 with low numbers, difficulties, and so on and so forth, could now be done very easily. In particular, mapping of genes on the chromosome. Because you went from recombination of the order of 10 -6 to the recombination of the order of 10 -1, that was an absolutely major event. And without this, Velma and Jacob would have not done their mapping work. So Hayes and Cavalli-Sforza who wrote another kind of HFr soon after were major contributors to the field. And so, the experiment we're going to see here is the experiment of interrupted mating. So, what is the principle of the interrupted mating? So, we're going to start with an HFr strain which is three union plus gau plus, and we're crossing this with an F-, which is 3 union minus gau minus. In addition, this strain is resistant to streptomycine, and this strain is sensitive to streptomycine. So you incubate the cell together, and at various time after incubation, you plate the cell on, for instance, a minimal medium, no threonine plus strep. So on this plate, the HFr will not grow because it's strep sensitive. The F- will not grow because it's threonine minus, only T+ recombinant. On this plate you will get only T+ recombinant. If you use another plate that has threonine present, but galactose instead of glucose, you will select the G+, Gal+ recombinant. So when you add the cell, that's times zero. You can take samples at different time and count the T+ recombinant. And this is this curve 1, this is T+. And curve 3 is G+. You see that as you wait, the number of recombinant goes up. There are more G+ than Gal+ recombinant, and you reach a plateau. Now, Jacob went to a meeting at Covetin Harbor a few years before that, and since he was a very devoted husband, he wanted to bring a gift to his wife. And so he'd seen the experiment that Hershey had done with phagian bacteria and the Hershey, the blender experiment, the whirring blender experiment. And so he'd say, but this is a useful cooking instrument. I can bring this to my wife. And so he bought one to put in his suitcase and gave it to his wife, who said who do you think you are? To bring me such an American junk, piece of junk, I'm cooking the French way and I don't use this kind of equipment. Remember we are in the early 50s. And so Jacob was a bit disappointed, and he put the whirring blender on a shelf in the kitchen, way up, so that it wouldn't irritate his wife. And then they said okay, but what happens when we do the conjugation of the HFr, with the F-? What if we could break the conjugation at different time? What would we see? And then he thought about the whirling blender that was taking dust in his home. So he took it, his wife was perfectly happy, and took it to the lab, and used the whirring blender for the second famous experiment with the whirring blender, the interruptive mating. And the interruptive mating involved the disruption mechanique, [FOREIGN] that's the way it's called. This was the result he obtained with the threonine. This is with a blender. The point with the blender is that if you interrupt the mating before ten minutes, you don't have recombinants. It takes ten minutes for the DNA to get in the cell. With galactose, it takes 30 minutes. This is very different because once you've interrupted, there is no chance that the conjugation will start again. So transfer of DNA is directional, the efficiency drops as you go longer and longer and longer. And then they also measured the threonine and galactose combination. So if you select for Gal+, most of the cells are T+. If you select for T+, only 25% of the cell are Gal+. There is something called asymmetry of recombination. So, they used this as a tool. They used more markers. And they selected on this plate, they selected for T+ streptomycine. And they selected 100, 200, 500 colonies. And at each time point, they measured how many were, for instance, azide-resistant, T1 resistant, Lac+, Gal+. And in this experiment, they found that it takes about 30 minutes for Gal to enter, only 20 minutes for Lac, 10 minutes for T1 and azide. So you can derive a map whose units are not miles, centimeters, nautic miles, or whatever metric, or whatever physical unit of distance. But when you use as a distance the minutes, the time is the unit of genetic linkage, genetic distance. So if you draw the map, you would draw the map by saying that at zero time, you have the entry point. At 9.5 minutes, you have azide, at 10 minutes you have T1, at 20 minutes, you have Lac, at 30 minutes, you have Gal. This is the beginning of the map that within a few years, would become the circular map of e-coli genome with 100 minutes of DNA. The 100 minutes today is the time required to transfer 4 x 10 to the 6 base pair. So you transfer 4 million in 100 minutes. In one minute, you transfer 40,000 base pairs. It's not a very fast process. You transfer less than 1,000 base pair per second. Remember, replication goes ten times faster, so this is a slow process. And it's a process that can be interrupted by a whirring blender or by shaking or by anything you want.
