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Genetics 3 Biology > Introduction to Biology
Video Lectures - Lecture 8/nTopics covered: /nGenetics 3/nInstructor: /nProf. Eric Lander/nTranscript - Lecture 8/nLet's dive in today and look at how geneticists use genetics. I've told you up until now about some of the history of genetics and how it gave rise to our understandings about genetic transmission in traits, about genetic mapping, linkage analysis, how all this helped confirm the Chromosomes Theory./nAnd we wove in a number of concepts about how scientific theories are developed and data is interpreted and intuitions are made, and then how they're actually proven, what sort of evidence it takes to actually achieve conscientious around theories. And that often takes sometimes years, many times decades before full contentious is achieved around things. Today I want to turn a little bit to the experimental uses of genetics in a more day-to-day fashion. And you will recall this coat of arms that I put up here./nFunction. Gene. Protein. Biochemistry. Genetics. And I told you about how these were two different ways to study biological function. Today I want to talk a little bit about how we use genetics to study biological function. And, in particular, I'm going to pick some examples of how we use genetics to study biological function that have to do with the biological functions of biochemistry./nSo already we're beginning to look ahead to this connection between gene and protein, which molecular biology will establish for us. So, suppose you want to do genetics. You've got to study some organism. We talked already about Mendel's choice of organism, the pea./nWe talked about some of its advantages and disadvantages. Advantages you could get pure breeding strains in the market, you could, when you're done with the experiments, feed it to the other monks. There were a lot of things like that, that were advantageous about the pea, but it had problems of generation time. You would only get, certainly in Europe, a generation or so a year. In Northern Europe maybe you could squeeze a second generation in not so good./nFruit flies, a very attractive system in many respects because you could grow many, much larger numbers. The generation time is on the order of two weeks or so to go from a fertilized fly embryo, a fly egg developing into a fly, developing into a mature adult, able itself to have offspring. So, very attractive. There are other systems that people studied. And, of course, one of the reasons they study this system is because it's interesting, I'm sorry, because it's tractable. And the other reason is because it's interesting. So, tractability is very important to a geneticist, right?/nThe number of whale geneticists is few, for the most part. But we also want to choose our system because of what it will tell us about the system we want to study. Like if you want to study distinctive things about the immune system, you might want to study them in mice, or if you could even study them in people, although you can't set up crosses in people. We'll come to that on Monday. If you wanted to study things about basic aspects of development, you might study them in fruit flies./nAnd if you wanted to study basic biochemistry, the place to study basic biochemistry might best be done in single-celled organisms, which also have to carry out biochemical pathways like glycolysis and synthesis of amino acids and things like that. They're going to be, by far, the most tractable systems. And so, people are particularly fond for doing things like studying basic biochemistry and many other aspects of basic molecular biology to studying the organism yeast./nYeast is a friend of human beings. Certainly, yeast has been an intensely studied organism because of its practical benefits in the making of bread, in the making of beer. So, fermentation processes, dough rising and all that. But yeast also is a tremendously important organism for the geneticist. It is an extremely elegant experimental system. Yeast is a fungus./nIt is a single-celled eukaryote. That is true nucleus. It's got chromosomes that pair up. It's cells, through a first order approximation, that are an awful lot like your cells in terms of having all of the basic important eukaryotic organelles in the nucleus, mitochondria, other things like that./nSo, yeast is a great model for many purposes. And we're not going to talk much about the cell biology of yeast, but I do want to talk about the husbandry of yeast, how it is that you grow yeast. So, the way a geneticists grows yeast is take growth medium that has lots of rich nutrients. You could take a broth with lots of amino acids and all sorts of stuff, you know, a little bit of salt, lots of water of course./nAnd if you take a single yeast cell and it's got lots and lots of rich nutrients in this broth here, you put your yeast cell into the broth, so I will do that. Here's my flask, here's my little rod which has a yeast cell or a couple of yeast cells on the end of it. I put it in there and I grow it at an appropriate temperature. Let's say 30 degrees, for example, would be a nice temperature./nI could do that if I wanted to. Then a C obviously. I grow it up and I get a culture of yeast in there. And I can tell because this nice clear broth is now all cloudy with yeast that's grown up in it. Now I want to study these guys, so what I do is I pour them out onto a Petri plate. The Petri plate has on it a medium, a solid medium, an agar medium that again has nutrients./nAnd if I pour this out, and I pour out a lot of it, what will happen? Well, there will be yeast all over the place and it will be very smootsie. There will be like yeast cells everywhere and it's not very organized. So, what I want to do is I want to take that and I want to dilute it. I want to take only a little bit of the broth and put a little bit of the broth on my plate./nMaybe I'll have diluted it first. And then I want to spread it around with a little spreader, here's a little glass spreader maybe or something, and push it back and forth, so that really there are just individual single cells scattered randomly, scattered around. And so, then this cell begins to grow and divide and divide and divide and I get a colony./nA little hill of cells all of which descend from a single cell that was put into that position. And the reason I know that they all descend from a single cell is because most of this plate does not have cells on it. Most of the plate is sparse. I've just got cells, cells, cells, cells scattered about. And because of that I know that these had of been individual events. These things are called colonies./nNow, when yeast grows and divides like that, let's take a moment and talk about its life cycle. We'll introduce its life cycle here. Yeast proper eukaryote, so it has a diploid stage. It grows as a diploid. And it can undergo mitosis in which all of the chromosomes line up, as we talked about./nThey've already pre-replicated so that they'll be ready to divide up and give one to each daughter cell, and there you go. N for yeast is 16. Yeast happens to have 16 pairs of chromosomes. Peas had seven. Humans have 23 pairs. Every organism has its own yeast of 16./nNow, what we do is we undergo meiosis to make haploid cells, sperm or eggs in the human population. Yeast also undergoes meiosis to make spores. It sporulates and it produces spores. And it turns out these spores, of course as you would expect, have N chromosomes. They undergo meiosis just as we drew it on the board. And these can come in two flavors./nThey happen to come not in male and females, but A and alpha, there you go. A and alpha cells can mate together to produce, again, a diploid. They fertilize and can produce a diploid. They fuse to do that. And you now get back to a diploid from your haploid. So, this looks just identical to the human genetic cycle here, but there is one difference. What's the difference?/nSorry? Time. Yes, it's true. Yeast can divide much more rapidly. Yeast can have offspring extremely rapidly over a course of a day or so. And humans take somewhat longer than that. They, for example, have to wait until they get out of college to be able to reproduce mostly. What else? There's one other important thing. It turns out that yeast can also undergo mitosis as a haploid./nIn other words, the haploid cells of yeast, when it makes individual haploids, they can continue to grow indefinitely. By contrast, your gametes cannot. You do not have an independent human stage in which you are haploid, or your gametes are haploid./nWhereas, yeast can hang out as a haploid for a very long time until it decides it wants to mate. This is very convenient for geneticists. Geneticists like this because it means we can grow the thing as a diploid, we can grow the thing as a haploid. When we want to mate them, we can mate them together, but we can also study them alone. And, you could imagine, this is going to be really good for studying recessive traits, right? So, that's one of the reasons why geneticists are fond of yeast. There are many reasons geneticists are fond of yeast. Just growing yeast, it smells very nice in the lab./nFor example, try growing E. coli by comparison. So, now, it turns out that yeast is very happy if you grow it on rich medium. But yeast can grow on minimal media with very few macro molecules./nIt needs a carbon source which is some sugar that it can ferment. It needs a nitrogen. It needs some simple source of nitrogen. It needs some simple source of nitrogen. It needs a source of phosphorus. It needs some other trace salts and things like that./nAnd obviously it needs some water. That's it. If you think about what's in a yeast cell, like it's got phospholipid bilayers. But you're not giving it any phospholipids. Why is it able to grow? It makes them. What about proteins? They're made up of 20 amino acids. You're not giving it any amino acids. Why? It makes them./nYeast is extraordinarily self-reliant. You, by contrast, are not as self-reliant. There are a number of amino acids which, if I don't give you, you can't live because you don't actually have the ability to make those amino acids. But yeast is able to make the vast majority of things. Basically, you almost just needed to give it the elements. As for carbon sources and things like that, it's very happy with a wide variety of fermentable sugars. You can give it glucose. You can give it sucrose./nYou can give it galactose. You can give it fructose and it will deal. So, yeast is very well set up metabolically. So, it's got all of these pathways of the sort Bob has talked about for being able to breakdown the things you give it and being able to synthesize up the things it needs. Now, yeast, of course, is not stupid. Because if you give it amino acids it will use it. If you give it all sorts of other things it will use it./nSo, yeast is able to use rich media that have lots of complex nutrients and macromolecules. So, it has an ability, it has everything it needs to make these things, but it has an ability to regulate that./nSo, the processes, the enzymatic pathways that produce complex macromolecules, amino acids, phospholipids, et cetera, will be down regulated, shut off, or at least decreased if you provide it with these macromolecules. That's an interesting question of how it manages to regulate its biochemistry. Why does it care? Why doesn't it, why not just have those pathways be on all the time?/nSorry? Waste of energy. It needs ATP. It costs money. So, at the beginning probably they were on all the time, but some yeast evolves, or some precursor to yeast evolves that's able to regulate it. That one is able to be more frugal with its energy. It outgrows its other ones and then another, dah, dah, dah. Any place you can make a few ATPs here or there, eventually the organism that does it will out compete the organism that doesn't. And so, rather fine control of this, which is a topic we'll come to in a couple of days, gene regulation and other kinds of pathway regulation is very important./nOK. So, we want to know how does it do it? What are the enzymes? What are the pathways? How does it actually make, oh, I don't know, arginine? How does it make arginine, amino acid? How would you find out how yeast makes arginine? How can yeast synthesize arginines?/nSo, you remember our picture that the biochemist wants to study a problem by grinding up the cell and purifying a component able to do something. So, a biochemist might want to grind up the cell and purify an enzyme that can make arginine. Form what, of course, is an interesting question? And then the thing that made the thing that was used to substrate, et cetera, et cetera. What would a geneticist do? How does a geneticist approach the problem with how does yeast make arginine?/nFind a yeast that cannot make it, that's what we do. That is. So, what we need is a mutant. A geneticist wants a yeast that cannot make it. A geneticist wants mutants. How do you find the mutant? You find the mutant by going on a mutant hunt./nThat is what geneticists refer to it as. And it's a very exciting thing. You go off, load up the guns and go off into the bush on a mutant hunt. And so, I want to talk about the strategy for a mutant hunt. How do we look for a yeast that can't make arginine? Sorry? Cannot. I've got a yeast that can make arginine, because normal wild type yeast can grow on minimal media without arginine supplied. And, when I examine it, it's got arginine in it./nYes? So, who should I find? Proteins that contain arginine and then it doesn't have the proteins that doesn't have arginine. Interesting. Now, the problem is almost all proteins will have an arginine, or the vast majority of them. And a yeast that lacked all those proteins that didn't have arginine would not be much of a yeast. I think it would be pretty dead./nSo, it's a good thought if it was a more dispensable function. But that's going to be tough. Or, maybe I can use the fact that it's dead. Now in a sense, can I find the yeast? Yes? You had a thought on this. Yes? Kill all yeast that make arginine, excellent. So, if I had a chemical agent that could kill yeast that can make arginine, I could only get the yeast that make it. How would I do that?/nThat's a very interesting idea. You're right. You could construct the chemical molecule in the arginine pathway which when it was broken down enzymatically made some toxic product, and only those yeasts that couldn't break it down would be able to grow, et cetera, et cetera, and I could select. That's a very cleaver idea. But I'd also have to know an awful lot about the pathway in advance. So, suppose I didn't' know the pathway. Suppose I knew nothing about how arginine gets made. Yes?/nExcellent. So, I take, I mean geneticists are simpleminded folks and they like simple solutions. Take medium in which you've given the yeast arginine, grow it up, and then pour it out on a plate that doesn't have arginine. Everybody got this idea? So, we're going to take yeast./nWe're going to grow it up in medium which contains arginine with arginine. So, now yeasts, those mutants that arose by chance that are unable to make their own arginine are still able to grow here. And then we dump it out onto a plate that has minimal media without arginine, no arginine, and those ones that can grow up are the ones that we're not interested in./nAnd the ones that don't appear are the ones we're interested in. But, wait a second, that's the problem, isn't it, because they're not here. How do we study them if they're not there? What can we do about that? Yes. You want to see if you can help us. Remove the ones that grew up. So, get in there, scrap them off, now put some arginine on./nWe're getting to the idea. Maybe we can set this up more elegantly, though. Thoughts? How can we, yes? Make a bet? Make a guess? I can make that guess, but how do I find them?/nHere's a simple, simple, simple idea. Let me try a simple idea. How about I grow up these yeast, and instead of plating them on minimal medium, let's be good to them. Let's plate them on minimal medium. Good. That's interesting./nLet's plate them on minimal medium plus arginine. Or, actually, if we wanted to, we could even plate them on rich medium. We'll be really good to them. Either way. So, now, let's let each one grow up. And here will be the ones that can grow and the ones that can't grow with arginine./nNow let me take a plate that is minimal medium. And now let me take a toothpick, put a little toothpick there and carry over this colony to there. Let me take a toothpick and carry this guy over to here and a toothpick and carry this guy to here, and a toothpick, and a toothpick, and a toothpick. And all I have to do is keep transferring, one at a time, these colonies./nAnd now I can see that somewhere there was a colony that grew fine when I gave it, say, rich medium, or minimal plus arginine, and a colony that didn't grow when I put it on minimal medium. That would at least show, so, of course, the issue is I first have to find them by growing them on something where I've given the arginine and then I can see that they can't grow. All right. This is what geneticists basically do. What happens if I grew them on rich medium and I transferred them to minimal medium? Why might something not grow?/nIt might be missing the ability to make tryptophan. It might be missing the ability to make proline. It might be missing the ability to make something else. So, what I can do is, if I wanted to, make a very broad mutant hunt. I could just first grow on rich medium and then plate on minimal medium and any yeast that has lost the ability to make some essential nutrient will be evident by its absence on the minimal medium plate. So, we have for yeasts./nYeasts that are able to grow on minimal media are called prototrophs. They are the wild type that can grow on minimal media. They can make everything themselves. Yeasts that need help, that cannot grow by themselves, that need help, that need a supplement are called auxotrophs. Auxo obviously meaning help./nSo, it's a mutant that has lost the ability to grow on minimal medium and that it needs a supplement of some kind. So, if I wanted to, I could just first collect lots and lots and lots of auxotrophs and then figure out what they need. So, I might collect a large collection of auxotrophs./nAnd then test to see if supplying arginine rescues them. I could also test tryptophan. So, if I only, only, only cared about finding arginine auxotrophs, I could just grow them on minimal plus arginine and then test them on minimal./nAnd then I would know in advance, these guys all grew with arginine on minimal and didn't grow without arginine, and I'd know it was arginine. Or, if I was in an expansive mood, I could test them on rich medium, collect everybody who's unable to grow on minimal, and then work out what the reason is. Is it arginine? Is it proline? Is it whatever? And it depends how much work you're interested in doing and how complete the study is you want to do. Either way, we could end up with a collection of arginine auxotrophs./nOrganisms that are mutant for the ability to make their own arginine and require it to be supplied to them in the medium. All right. I might get, depending on how much work I'm willing to do, dozens of independent colonies unable to grow without arginine. I might get hundreds if I'm willing to do enough work. I can get as many as I want./nOur goal now is to study them and find out why they're unable to do that. I have a quick question? Those yeast cells we plated, where they haploid or diploid? We didn't say, did we? So, should they be haploid or diploid?/nHow many vote diploid? How many vote haploid? A lot of people vote haploid but aren't willing to express a reason why. Why haploid? Right. Excellent. Excellent, although genes are not recessive, but OK. A little detail. Phenotypes are recessive./nTell me a little more of what you're thinking about. We'll have it out later on this point, yes. So, suppose we were looking in a haploid. I take your point, even if on nomenclature I want to push back a bit. So, suppose it's a diploid and suppose we have now two copies of this chromosome here in the diploid./nAnd suppose there's a gene over here that encodes an enzyme that we now is necessary to make arginine, or that somebody knows is necessary to make arginine. Let's image that that's the case. In order to get haploid yeast that is unable to make arginine due to a mutation in this gene, you need to have some kind of a mutation in this copy. What about in the diploid yeast? In order to make this yeast unable to grow without arginine, do we need a mutation in both copies?/nWell, the answer is probably. The truth is actually a bit more complicated, but let's suppose it was the case that even one copy of the functional gene was sufficient to carry out the enzymatic step, then the answer would be yeah, we'd need a mutation of both copies. What's the chance of finding a yeast that has a mutation in both copies? It's obviously much less than the chance of finding a yeast that had a mutation of one copy./nSo, we're much better to go searching in the haploid where the phenotype will be revealed much more easily by virtue of just the single mutation rather than having to, by chance, encounter one that had mutations in both copies. Now, the reason I'm a little bit cautious here is because notwithstanding the textbooks, it's not always the case that everything like this is a recessive trait. It's possible that auxotrophy for arginine could be a dominant trait./nSo, how could that be? Well, auxotrophy could be a recessive trait. Suppose there's some enzymatic pathway, A goes to B goes to C goes to D, and this encodes an enzyme that carries out a particular biochemical step. Well, if the gene is broken, if the gene is missing, if the gene doesn't make the protein, as you guys all know that that's what happens, then you don't have the enzyme, you can't do the pathway. And it is usually the case that having just one copy is sufficient./nBecause having a little bit of enzyme the pathway may work slower but it will still work just fine and you'll eventually get arginine made. But it's occasionally possible, I note since you guys are sophisticated, that sometimes a gene can encode a protein which not only doesn't work but screws up the other working copies of the protein. Suppose the enzyme that did this were a tetramer. It had several subunits that had come together./nA mutant copy of an enzyme, when it forms into a tetramer, might somehow disrupt all the other good copies that are around. And that does happen sometimes. It can happen that you're going to have an inability to make your own arginine be a dominantly inherited trait. So, you actually have to test whether it's recessive or dominant. Often it will be recessive. So, usually most of these simple auxotrophs are recessive traits./nOccasionally some are dominant. So, now, suppose we get a whole collection of Arg auxotrophs, and we'll just give them a name. I don't know. Here's my collection. We'll call the first one, for lack of anything terribly creative, Arg 1, Arg 2, Arg 3, et cetera, each being an individual strain from growing up originally for a single colony that is unable to produce its own arginine./nWe now want to take this collection and characterize it. How many distinct genes does this affect? Are these mutants perhaps all in the same gene? Are they in a hundred different genes? How could we tell?/nNow, of course, if you're a biochemist, you already know the protein you can see and dah, dah, dah. But, if you know the answer, well, why are asking then, right? A geneticist goes out to ask this question because he or she wants to know all the possible ways you can disrupt the cell so it cannot make arginine. And we don't know in advance what those ways are, so how are we going to be able to tell whether or not different mutations affect the same gene, the same function in yeast? It's an interesting question./nGeneticists do a variety of tests. The first test that a geneticist does to characterize a mutant is by tests of recessivity or dominance, whichever way you want to put it./nWe want to take each mutant and test whether it is recessive or dominant as a phenotype, whether the phenotype, the auxotrophy for arginine is recessive or dominant. So, here's mutant number one, the mutant cell carrying this mutation here. Conceptually it affects some gene. I'm going to label it Arg 1. We don't know where it is in the genome. There are other chromosomes here as well. Here's my mutant cell. How am I going to find out whether or not the auxotrophy for arginine is recessive or dominant?/nYup? With what? Cross it with a haploid that is a prototroph, or I could just say cross it with wild type, right? Perfect. So, make a cross here, very good, with wild type plus there. How do I know it's plus there? This is wild type. Wild type is defined as the normal form./nAnd so, because I said this is what we're using as wild type, it's necessarily plus because we're measuring mutations relative to wild type. So, what happens when we get here? We now, when we cross we get a diploid, and Arg 1 plus. Now, how do we know whether or not that phenotype was recessive or dominant? Sorry? It's what shows up when we try to grow it./nSo, when we cross it, what kind of plate should we grow it on first? Should we grow it on minimal or rich? We better grow it on rich because just in case it doesn't, it can't make its own arginine, we better first let it grow and then test it. So, let's grow it on rich medium. We'll cross these together, grow it on rich medium. So, grow on rich, test on minimal./nOK? And we'll be able to check out the phenotype as to whether or not the phenotype is wild type or mutant. All right. So, we could do that. And we'll test the first one and the second one and third one and the fourth one. And, for each of these, we'll write down whether it's recessive or a dominant auxotroph. Now, let me assume that all the ones we're talking about are recessive phenotypes./nBecause everything I'm about to say is very much harder if it turned out any of them were dominant. So, we're going to assume. Let's assume now, but it's not always the case, we'll assume that the collection, maybe Arg 100, are all recessive auxotrophies, the phenotype is recessive./nNow, how do I tell if they're in the same gene or not? So, now I want to characterize my mutant by some other test that will tell me whether or not Arg 1 and Arg 2 are in the same gene./nSuppose Arg 1 and Arg 2 are in different genes. Cross them. What will happen? Right. So, to repeat that, if I cross together the two mutants and they're in different genes, each will have at least, the each will be contributing a good copy, a functional copy, a wild type copy of one of the genes./nSo, let's walk this through. Interesting. Interesting. So, suppose I take a situation where I've got Arg 1, a mutation in a gene over here, on this chromosome, and on the other chromosome I've got a wild type copy. My Arg 1 mutant is mutated in a gene here./nI've got this other gene here, which is normal. And I'm going to cross that now by the strain that has a wild type copy here for this first gene, but it has a mutation in the second gene. When I cross them together, I now get me a diploid cell here, which is Arg 1, a mutation there, plus there, plus copy here, and Arg 2./nWill having one copy, one working copy of this gene be enough to make the enzyme? No? In other words, is the wild type phenotype dominant to this auxotrophy, or is the auxotrophy attributable to this gene recessive?/nYes. Why? Because we assumed it. Why did we assume it? So I would be able to say this, right? OK. If it wasn't we'd be in trouble. But by assuming that we're working with a recessive phenotype, then we know that this will be enough to save the yeast. What about here? Enough to save the yeast so it will grow without arginine./nBy contrast, suppose it was the case that this cell here, Arg 1, and suppose our other mutant that we had isolated in our mutant hunt was a mutation Arg 2 in the same gene. Suppose these were the same gene. When I cross them together I now have a cell that is Arg 1, Arg 2./nIn other words, its genotype is Arg 1 over Arg 2, name of mutation. And can it grow? No growth without arginine. By contrast, the genotype here is Arg 1 over plus, plus over Arg 2./nI could even write Arg 2 over plus, but I just did that to indicate the chromosomes that they came from. All right. This is called a Test of Complementation because these two genes are able to compliment each other's defect./nIf two mutations compliment each other's defect then they are in different genes. OK? Boy, that's a noisy one./nSo, we're able to make a Complementation Table. Suppose I take a bunch of yeasts, wild type, WT, mutant number one, mutant number two, mutant number three, mutant number four. And suppose I cross them with each other in all pair-wise combinations./nI've assumed that all of these arginine auxotrophs have a recessive phenotype here. These are all my Arg mutants, and I'm assuming that this is recessive. What happens when I cross them and I test to see whether they can grow without arginine? If I cross wild type by wild type, can it grow without arginine? Yeah. Normal phenotype. So, plus is going to mean prototrophic./nMinus will mean auxotrophic for arginine. What happens when I cross wild type with mutant number one? It grows. Why? By assumption, these were all recessive. I'm only testing recessive ones. Two. Three. Four. When I cross in this direction, wild type by these guys. This is going to be a symmetric matrix, of course, right? OK. Now, what happens when I cross mutant one by mutant one?/nI now have a diploid. Will it be able to grow without arginine? No. Why not? It has no working copies of that gene, so I'm going to put a minus there. What about mutant two with mutant two? Minus. What about mutant three with mutant three? Minus. What about mutant four with mutant four? Minus. Now, what happens when I cross mutant one by mutant two? It depends. It might be plus or might be minus./nIf they're in the same gene, minus. Different genes, could be plus. So, here's some data. So, all this is compelled. But the kind of data, ooh, I'll use a color. Isn't that fun? They want me to use colors over there. Here we go. Suppose the data were minus, minus, plus, plus, plus, plus, minus, minus, plus, plus, plus, plus./nWhat would it be? What conclusion could we draw? Is mutant one and mutant three in the same gene? They complement each other? No. But is one in the same gene as two? Yes. In fact, this box and this box here define the genes beautifully./nThe groups that failed to complement define mutations in the same gene. These are called Complementation Groups because they don't compliment, OK? It's a little complicated but that's all right. These are called Complementation Groups because all the members of the complementation group, namely Arg 1 and Arg 2, failed to compliment each other. They could be called Failure To Compliment Groups, but it would be too long./nOK? So, there you go. You can take hundreds of mutants and organize them into complementation groups and thereby know which ones go to the same gene. And now, if I want to study the genes, I only have to study the distinct complementation groups. Last thing, which we'll just have time to do, are what's called tests of epistasis. We'll probably run just a moment or two over on this./nSuppose a biochemist were collaborating with a geneticist and had studied what he or she thought was the pathway for making arginine. Some precursor alpha goes to precursor beta, goes to precursor gamma, goes to arginine. And suppose specific genes were needed to encode specific proteins./nI'll call them Arg A, Arg B, Arg C to catalyze each step of this biochemical reaction. The geneticist and the biochemist could collaborate with each other to study whether these mutants, these particular genes now that had been identified, affected each step of the pathway. And here's how they might do it./nThey might take wild type yeast, mutant, well, they wouldn't know in advance whether or not it was missing the ability to grow on each of, whether it was missing each of these enzymes, but let's think conceptually. Suppose we had a mutant that was, a strain that was wild type, Arg A minus, Arg B, minus, Arg C minus, unable to make this enzyme, this enzyme, this enzyme. And suppose we helped it along. Suppose we gave the mutant arginine. Suppose we supplement and grow it on media with arginine./nWhich ones will be able to grow with arginine? Can wild type grow if it's given arginine? What about Arg A minus? B minus? C minus? What if instead we offer it precursor gamma? Will wild type be able to grow if it's given precursor gamma? Sure. What about Arg A minus? No, because it still is stuck at this step./nIt cannot. What about Arg B minus? What about Arg C minus? Really? It hasn't got this enzyme. What's it going to do with gamma? It ain't got anything to do with gamma, no enzyme. Suppose I gave it beta. Wild type, can it grow? What about Arg A minus? No, because it can go from alpha to beta, but it can't go to gamma./nIt cannot grow. What about Arg B minus? I've given it beta, but it can't do anything with beta because it hasn't got this gene. What about Arg C minus? Wait a second. What did I just do? We're just backward. Sorry. If we gave it gamma, I just got lost here./nIf we gave it gamma it was able to grow, well, we are completely wrong, guys. It's able to grow here. Thank you. Let's go back on that. You should have caught me before. My mistake. If we have it gamma it's able to, if it's a mutant here it can grow because it bypasses this problem. And having gamma is enough. If I gave it beta, sorry, if I gave it gamma and its mutation was here it can grow. Sorry./nNow, if I gave it here beta, and its mutation was here, it can still grow, right? But if its mutation is here it can't and if its mutation is here it can't. That's better. I was getting worried there for a while myself. Suppose I gave it alpha. Wild type can grow. If I give this guy alpha, will that help if he's mutant in A? No. Can it help if he's mutant in B? No./nCan it help if he's mutant in C? No. Sorry. There we go. I usually start at the other end of this picture. So, what you can see is these mutants have different phenotypes with respect to being able to supplement them with different chemicals. Now, let me ask in our last two minutes, I'll run two minutes over here. Suppose I gave you a mutant that was a double homozygote./nSuppose it was Arg B minus, Arg B minus, sorry, Arg B minus and Arg C minus. Suppose it was a double mutant, it lacked both this and this. Which line of my table would it resemble? Would it look like the first line, the second line or the third line of my table?/nSecond line. Why's that? If I'm lacking B, I'm already in trouble here. And also lacking C doesn't matter. So, I will look, just like a mutant who lacks B. So, in other words, I'm able, if I know something about the biochemistry of a pathway and I can break my arginine mutants up into different kinds of phenotypes here by their response to different steps in a pathway, I can then look at combinations of mutants./nAnd I can say if I have a double mutant missing both B and C, does it look like B or does it look C when I put them together? And it turns out that if it looks like B then B was further upstream in the pathway./nSo, it turns out that geneticists and biochemists can collaborate based on the phenotype of the organism sometimes to infer aspects of the biochemical pathway. These are the kinds of things a geneticist does to be able to characterize mutants on a mutant hunt. Next time what I want to do is talk about characterizing mutants in a very different kind of organism, namely the human being.
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Genetics 1 Biology > Introduction to Biology
Video Lectures - Lecture 6/nTopics covered: /nGenetics 1/nInstructor: /nProf. Eric Lander/nTranscript - Lecture 6/nI am the other half of the teaching team for 7.01. You've already gotten to meet my good colleague Bob Weinberg. My name is Eric Lander. And Bob and I are both faculty here in the Biology Department. In fact, we're both members over at the Whitehead Institute for Biomedical Research. In fact, we just spent the whole weekend together at the Whitehead Retreat. And so, Bob and I have been doing this course together for a number of years. And we very much love it. I am --/nI'll take a brief moment and introduce myself, since I haven't had the opportunity to do so yet. I am by training, well, actually, I'm really a geneticist. By training I'm actually a pure mathematician. That was actually what my undergraduate degree was in, and even my PhD was in, but then wandered into biology. And for the last almost 20 years, I have been doing genetics in some form or another. So I love genetics and look forward to talking a lot about genetics. And it's really lovely that my first lecture today is actually going to be our first introduction to genetics. I am --/nJust for other backgrounds, I direct this new Broad Institute that is here. And it's actually a joint institute between MIT and Harvard. And you will know it now as a hole in the ground next to Legal Seafood. If you see a bunch of cranes and things opposite the biology building and opposite Legal Seafood next to the Whitehead, that's the Broad Institute. And we have ambition some day to be more than the hole in the ground but to actually rise above the ground. And the Broad is about genomic medicine and using genomes and things like that./nAnd the Broad Institute includes this center at MIT that was one of the leading participants in the Human Genome project. So that's a lot of what I do with my day job, in addition to teaching, is work on things like the Human Genome project. And, now that we actually have a sequence to the human genome, figuring out what in the world it all means. And I hope I'll get a chance to tell you, during the course of this class, about the human genome and about what's in it and things like that./nLike I say, that's one of the things I tremendously love about teaching biology as opposed, if I can get in trouble, to any of the other required introductory courses, is that our curriculum changes every year because the field is moving so rapidly. I look back at what we taught ten years ago in this course, because I've been teaching it that long, and all sorts of open questions now we know the answers to and are part of the curriculum. Some of the things we thought we knew we now know are false and we know new things. And every year we get to introduce new stuff./nAnd I know, I mean with all due respect to calculus, it's just not the case for calculus that there's anything really new to introduce. Most of it sort of settled down about three or four centuries ago. And, you know, that's just not the case with what we do. Anyway, so that's why I love it. All right. So Bob has been talking to you about biochemistry largely./nAnd I'm going to now turn to genetics. But I want you to understand that that is an overarching framework that explains how all the materials you're going to see, at least in the first half or more of this course fit together. And Bob may have mentioned it, but I'm going to mention it again, I would use this following diagram as kind of our roadmap or subway map of where we're going in this course./nWhat we really want to do is understand biological function. That's what we most want. How is it that an organism is able to breathe in air and distribute it to its cells? How is it that an organism is able to move its muscles? How is it that an organism is able to fight off invaders to its body, microbes, things like that? How is it that an embryo develops into a full adult? Zillions of questions. That's what I mean by biological function. The two complimentary approaches to studying biological function, over the course of the past century or so in biology, have been the following./nThere have been the biochemists. Biochemistry seeks to break down the organism into individual components and study them on their own in a test tube. They will take an organisms, and to a biochemist wishing to study the beauty of a butterfly flapping in the wind and understanding all of the mechanics of how it could possibly flap those wings and all, he or she would start by taking the butterfly, putting it in the blender, pressing puree and making an extract, and trying to purify individual components that would explain muscles moving back and forth and all that./nThis is, of course, a geneticist's point of view, but it's all right. You have Bob who will represent biochemistry just fine. And they want to purify out individual components. Individual components away from the organism./nAnd the most important individual type of component that they study are proteins because there are zillions of proteins and they do all sorts of things in the body. And so you could say, in some sense, that this whole theme of biochemistry, which got started at the turn of the 20th century, really just a few years before the turn of the 20th century, of grinding up an organism, studying its components and being able to find, for example, I want to understand how I can digest lunch. Well./nOr how yeast can digest the sugar. Grind up yeast, fractionate it and find some protein that's able to digest the sugar all by itself without the rest of the organism, an enzyme to do that. That's the logic of biochemistry. Genetics is the complimentary point of view. Genetics is the study of organisms minus one component. Of course, what I mean by that are mutants./nThe geneticist who wants to understand the butterflies and how the butterfly can fly would isolate butterfly strains that have lost the ability to fly. And ideally one is extremely closely related to the normal butterfly, but for some reason, ideally due to the mutation of a single component they're now unable to fly. And the geneticist would then say, ah-ha, that component must matter an awful lot for the ability to fly because the butterfly that lacks that component cannot fly. It's a totally complimentary point of view./nAnd the objects the geneticists study in order to do that are genes. Now, what is of course hard for you guys to understand but will form a structure for some of the lectures that I'm going to give over the continuing part of this course, is that through most of the 20th century the folks who studied biochemistry and tried to understand proteins and the folks who studied genetics and tried to understand mutants had nothing to say to each other. They didn't speak the same language./nThey had nothing to relate to each other by because there was no idea of how this gene stuff, which started as a totally abstract business, could possibly relate to this protein stuff which started as a very practical in the test-tube thing. And they went for a very long time as if they were just ships sailing in the dark unaware of each other. And I exaggerate, but it's more true than not. The great intellectual event was the unification of these two points of view through the discipline of molecular biology./nMolecular biology was the discipline that realized, oh, my goodness, these are two different sides of the same coin. That, in fact, genes encode proteins, proteins are encoded by genes. Ah-ha. This was a wonderful and important thunder clap in the 20th century. Now, it was a theoretical piece of information at first./nThe idea that genes and proteins were related in this way was abstract, very important, but you couldn't do anything really with it, because it turned out you couldn't actually work with individual genes. The next great revolution of the 20th century was a technological revolution that let you actually work with genes./nAnd that was the recombinant DNA revolution in which the tools to be able to study genes on their own away from the organism, study proteins, use genes to figure out what protein they encode, given a protein and figure out what the gene is, given a gene and actually go in and make a mutant in it, not wait for a random one to rise in the lab but deliberately knock it out, all of that operationalized this intellectual procedure, this intellectual framework. So that is, in some sense, a roadmap to coming lectures that I'm going to give./nI'm going to talk about genetics, I'm going to talk about molecular biology, and I'm going to talk about recombinant DNA. That's the structure of the next several weeks of this course. And what I want you to do is to recognize that although we're going to dive down into the individual components of it, everything we're going to do over the coming weeks fits into this very amazing intellectual framework. And this is the intellectual framework that you inherit as the new students coming into this field and going into the 21st century is all this was worked out in the last century./nYou now have an understanding of how all these pieces fit together, or at least you will, how these can be used to study biological function and, as I will also talk about, the recombinant DNA has grown into a world of genomics that has given us the complete picture of all of the components. It's actually not bad. You were very wise to have shown up when you did because an awful lot of that groundwork has now been laid. You know, if you would have come along 50 years earlier, you know, all that would have been slogged through. Right now you have this laid out for you very nicely. And that's sort of what the theme will be. OK?/nI would ask are there any questions, but there should be a zillion questions about that. This is just intended as a framework there. So let's now dive in. Section 1. And I'll give a bit more background today than I will in some of the other lectures, but we've got to get going. What I really want to do first is talk about, in fact, most of today will be about Mendel./nI confess, Mendel is my hero. He is one of my absolute heroes in science. I just love Mendel. And so I'll dwell on him a little bit today. Now, here's the problem with trying to tell you about Mendel. You already know about Mendel, right? Who here hasn't met Mendel and the peas and the stuff and all that in their high school textbooks? So what am I doing talking about Mendel today? Well, I think what you learn about Mendel in the textbooks in high school does not really bring out what really went on with Mendel's thinking, what's really important about those experiments, what's really interesting./nAnd so I want to ask you to put aside what you think you know about Mendel and let's go back over the setting of who Mendel was, what he was doing, how it all adds up. Because I think in Mendel you can find just the seeds of how to do great science. Now, for starters let me clear up, I'll take five minutes to clear up, four minutes to clear up some misconceptions about Mendel./nIt has generally been written that Mendel was this monk working in this monastery often in the Chez Republic, at that point in the Austro-Hungarian Empire, and he was isolated, working by himself, and it was amazing he discovered all this stuff. It's nonsense. Mendel working on genetics was no accident. It was the result of extraordinary historical and economic forces over the course of about three centuries that culminated Mendel./nLet me briefly explain why. It starts with the Age of Exploration. Europe starts sending out boats around the world, explorers to meet other parts of the world in the 1500s. The boats come back. They bring back stories of amazing lands. They also bring back odd plants, odd animals. People begin to look at these plants and animals. They begin to cross them, grow them and cross them, and look at the weird odd combinations of things that are going on./nAnd they say, wow, there's so much more variation out in the world than we thought about. Some of it's kind of useful. We can make new kinds of varieties of plants different than we had before, new kinds of varieties of apples. Now, it turns out that's not just an intellectual curiosity that that was the case because economics was changing in the face of Europe in the 1600s and in the 1700s with better transportation networks. So if you happen to be able to make a better apple, it was good, not just for your family, but you would be able to project that through lines of distribution to larger markets./nIt became economically sensible to invest your efforts in producing a better crop because you could sell it to more people because unified markets and transportation systems were developing across Europe. And, therefore, economic forces began to work toward getting a hold on the understanding of how you could do better breeding. Now, this turned out to be particularly important to the folks in Central Europe in the Austro-Hungarian Empire, which was the center of the textile industry./nThey were particularly concerned, in the late 1700s, about the fact that as the center of the textile industry they had to be concerned about the raw materials like wool that they used. Wool you could get from Central Europe, the Spanish had begun producing by breeding better sheep with better wool. This freaked out the guys in the Austro-Hungarian Empire because they were risking now losing this stuff to the Spanish because of their better sheep./nAnd they began, around 1800, to say we better start understanding how to do breeding. They put together societies to understand better the science of inheritance and breeding. By 1820, a society which was not about sheep but about plants, in fact, apples and grapes, the Pomological and Enological Society of Braunau was organized. Braunau being the capital of the Austro-Hungarian Empire./nAnd this society got all the town fathers of Braunau together. In those days it was just fathers, you know. Together in Braunau and started this society to encourage the scientific study of agricultural inheritance. They had this big dinner and they were drinking and things, and the speech is actually written down where the president gets up and says, "Some day the world may be as indebted as it is to Isaac Newton for physics. They may be as indebted to the City of Braunau for its contributions to inheritance."/nWhich is just eerie to read that in 1820 in setting up this society. That was their high hopes for what they would do. In particular, the president of this society, one CF Nap was president of the society as a side job, his main job was he was head of the Augustinian monastery in Braunau. So he began keeping an eye out for bright young math and physic students. Basically, you know, MIT kids coming out of high schools. And he identified a bunch of smart ones and attracted them to the monastery and gave them problems to work on./nHe particularly was impressed with this relatively poor kid, Gregor Mendel, who had been floundering around with a couple of things, didn't have bright family prospects, and attracted him to the monastery to work on problems of inheritance. So this was no accident. This was a biotech incubator that had been set up in the Austro-Hungarian Empire. Not of the sort we'd recognize today, but it's just fascinating to realize Mendel was not in a vacuum at all. He knew what he was doing here./nHe really wanted, for the good of mankind, to understand how to improve inheritance. But why do we celebrate Mendel today? We celebrate Mendel today because he went about it, lots of people were interested in this problem, right? You could probably find hundreds of people who tried to do something on this problem. Mendel was different because he went about it as a scientist. He went about it with a rigor and a persistence unlike all of his peers at the time./nSo let's think about what it was that Mendel did. So, anyway, forgive me for the historical digression, but I think it's interesting. What did Mendel do? Mendel started by taking peas. Now, he went off to the market and he got different varieties of peas. And he brought back all of these varieties of peas and he tried growing them. Now, actually, although I don't have the records, I'm sure he did lots more than peas./nHe brought probably lots of stuff and he tried growing it. And the first order of question he wanted to ask is if I study inheritance, I've got to start with something that has constant properties. This seems obvious to you guys, but it was not at all obvious at the time that the most important thing you could do, if you wanted to understand the transmission of traits and crosses and inheritance and all that, is not to set up any crosses. It was first to set up your experimental system and make sure it was rock solid./nHe probably devoted years to getting varieties of different plants, and in particular settling on peas, with a property that when he had peas with different traits, like whether or not the pea seed was round or wrinkled, which will be some of our favorite traits here, that when you simply selfed this plant, crossed it to itself and looked at the next generation, it bred true./nHard to emphasize how important that was, but this was careful experimental design./nSo many biological projects fail because people don't take the trouble to set up a system that's rock solid. They set up a system that's noisy and you're not really sure you're going to be able to interpret the data, etc. So Mendel did that. Very good./nAlways, no matter how long you continued to breed these things, you continued to get round or you continued to get wrinkled. Now Mendel was ready. He was ready to set up his first controlled cross./nSo what he did was he took a round pea and a wrinkled pea and he crossed them together./nNow, that's again some serious work. You first have to go along to one of the peas, cut off its little pollen producing organs so it doesn't self-fertilize because peas will self-fertilize. You've got to cut them off early, make sure it doesn't get its own pollen on it. Then you go over to the other one with a paint brush, you get some pollen and you paint the pollen on the first plant. That's how you set up the cross. If you screw it up you could have self-fertilization or the wind could carry some pollen from something from somewhere else. So it had to be done very carefully. He set it up./nAnd his first big-time observation was? Now, again, I know you know all this, so feel free to chime in. In the next generation all the peas were round. We denote generations with an F. F stands for filial meaning children. We sometimes denote them with a G for generation. Anyway, I tend to use F, and most geneticists tend to use F./nThe parental generation here is called F0, the first generation is called F1, the second generation F2, etc. So why was this a big deal? This was a huge big deal. If you took a poll, a CNN Gallup poll of Braunau at that time and you ask voters what do you think would happen if I cross a round pea to a wrinkled pea, what do you think the majority of voters would say? Well, maybe half and half or maybe all a little wrinkled, you know, a little puckered or something like that./nThe notion that one trait would be totally dominant over the other trait was by no means the general thinking. And you know what? It wasn't even the general case. If you took plants, you guys must know. If you take plants and you cross them, the F1 usually looks like some kind of a mix. It's some kind of a blend between the two. And, of course, that's because you're really looking at situations where you're crossing things in which zillions of different traits are being inherited and it's a hodgepodge./nBut Mendel had a situation here where he got an absolutely crisp dominance of one trait over the other. And so wrinkled completely disappears, round dominates, wrinkled disappears completely./nNow, next he does another generation./nHe goes to the second generation. And here he does this by selfing this plant. That is he simply kind of puts a bag over it and lets its own pollen fertilize itself or he takes a little brush and he brushes its own pollen onto it. And in the next generation his remarkable thing was he saw some rounds and some wrinkles./nWhat was remarkable about that? Wrinkled came back. I thought wrinkled was gone. And it didn't come back in some half-hearted way like a little puckered. It came back fully, totally, every bit as wrinkled as the parental wrinkled. And the rounds were every bit as round./nThese were discrete traits. Wrinkled reappeared, and it reappeared with no loss. No change in the phenotype, no change in the appearance. And that was very important because at the time some of the predominant models were blending of traits. And you would never imagine, if I were to take grape juice and water and blend them together to get some kind of pinkish thing that I would be able to separate that back out into clear water and deep dark grape juice./nBut somehow this trait had appeared. Thus, the trait was discrete. Big difference. Big news. This trait could be found still lurking there. It was merely hidden in the first generation. Mendel did one other thing, dear to my heart as someone trained as a mathematician, he counted. When he counted up the rounds and the wrinkles he found what?/nSorry? Three to one round to wrinkled? No, it's not. He found 5,474 to 1,850. That's what he found. Now, what do you recognize about that?/nThree to one? No, it's not. It's 2.96 to one. It's not three to one. What's this three to one business? [LAUGHTER] Why isn't there a famous 2.96 to one rule? No, no, I'm serious. Mendel did one more thing. He counted./nAnd then he did something a little bit outrageous. He intuited. He said although the data do not say three to one, notwithstanding your textbook, I think the data are trying to tell me it's three to one. [LAUGHTER] This is part of science. I'm sorry? Two sig figs, right. You know, this is actually a big deal because so many people are unwilling to kind of look at their data to say what's the data trying to tell me?/nAnd, of course, there are so many people who are too willing to look at their data and say what's the data trying to tell me? Because you can go off the tracks in both directions. So Mendel tried some experiments, 3.04 to one, 2.91 to one, etc. And occasionally, yes? No. How could he? Nobody had done this. He had no textbooks he could consult. So do you think it's possible he experimented with other things that didn't show these properties and said maybe these are lousy traits to work on./nI'm getting such good results on wrinkled, let's stay with wrinkled for a while. That is an incredible act of experimental judgment to know that some problems are too complicated, we'll come back to them later. It's not cheating. You get to say this is an interesting problem, I'm going to work on it. Not only that. I'll tell you, occasionally Mendel did these experiments and he got completely abhorrent results. They didn't match three to one at all. You know what he did? He threw out the data. Do you know why?/nNo, not small sample. Large numbers. He's sitting there in this garden. You know, I've actually been to Mendel's monastery. He's in the garden in Braunau. Remember, he's got to go cut off the little pollen producing organs, he's got to paint the stuff. What if he screws up? What if the wind blows and stuff like that? If an experiment was way off, he had to consider the possibility that he just screwed up because he hadn't gotten to it soon enough and pollen had blown in and had fertilized his plants. Now, boy, that's a dangerous thing to do, discarding data. But let's be honest./nSometimes experiments screw up. And if an experimentalist hasn't got enough judgment to know that sometimes you cannot believe the data you also can go wrong. So Mendel, who sometimes is accused for cheating for that, it's not at all cheating. What you have to do is say, OK, I've got a problem here. I'm going to redo this experiment a bunch more times. I'm always getting about this three to one thing, but occasionally I get something that's way off there and I feel comfortable saying that's an error. You can go wrong with that, but Mendel exercised very good judgment in excluding that rather than trying to muck this all up by saying occasionally I get something weird./nSo I know the textbook summarizes this beautiful 3:1 ratio, but so much creativity. First discipline of counting and creativity of interpretation went into all of this. So in the modern world what would Mendel do? In the modern world, upon seeing this three to one result which he, I will note, he saw for a couple of other traits. Actually, what he did next was he wanted to explain --/nThis was also part of his brilliance. He made a model, the model of what was going on. Mendel said how can I possibly explain this beautiful observation that for round and wrinkled, and for other traits, I observe an approximately 3:1 ratio in the F0, F1 and F2 generations? Mendel, my heart beats for Mendel. Oh./nA mathematician he is. He says let's make a very simple model. Let's assume that there are two factors of the control inheritance of this trait. I'll call them big R and little R. The round plants have big R and big R. They have two copies of this factor that controls shape. The wrinkled plant has two copies of the factor that control shape, and the copy of the factor they get is different. So the flavor here is big R, the flavor here is little R. This has two copies, this has two copies./nAnd let's assume that this plant transmits one at random of its two factors onto the next generation. It will transmit a big R. Let's assume that this transmits one of the two at random. It will transmit a little R. And that plant there in the middle will be big R over little R. And what will big R over little R be as an appearance? How does he know that that's going to be round?/nSorry? From the result. He knows because that's what happened. It's not an overwhelming reason. But to make the data work he's got to say, well, then this must be round. OK? So no points for that. He's just fitting the data. Then here, when you self this, the two parental gametes transmit either a big R, so we'll put it over here, big R, big R, little R, little R, they transmit./nAnd the offspring are of that type. You can either get that. Question there?/nCould be. So he had some knowledge./nBut, of course, this is his model. He's entitled to make his model. And you're saying he had good reasons to think in these. So everybody knows Mendel's model, right? So, now, in the modern world, the minute you've got data like this and you've got a model to explain it, what do you do? Publish. So Mendel, let's put Mendel as a young assistant professor who is all fired up about these results, writes this up for publication in Nature. It's a short thousand word letter to Nature, let's say. And he races it off, he emails it to the offices of Nature in London./nBecause he's in Europe, he'll use the London office of Nature saying I have this amazing result, I did these crosses. Here are the results here. And I have a model that explains the data perfectly. What does Nature do? Sorry? Why does it reject it? Well, the first thing he does is sends it out to referees, right? The way that scientific publication works is it chooses two or three anonymous referees. It sends the paper out anonymously to those two or three referees for comment saying we've received this interesting paper from this young monk in Austria. What do you think about it? Give us your opinions? Please write back in two weeks, etc./nSo you're the referees. You get Mendel's paper. What do you advise Nature? Publish or not? No. Why not? It's outrageous. Why? It's never been heard of. Yeah, that's great. But, I mean, you sound like a very conservative, you know, you cannot write that. You cannot say it's wrong because it's never been heard of. Yeah? Regenerate. It would be wonderful if referees could regenerate the result themselves, but it's not practical. For one thing, it takes a long time to grow peas./nThey might not have those strains of peas. The best test really would be independent replication of this, but unfortunately you cannot get the referee to reproduce each result before accepting the paper. So you have to go on the own internal results of the paper. Has Mendel proved his case for this model? How many people vote Mendel has proved his case for the model? He's my hero. How many people vote that he hasn't proved the case? How many people are conscience abstainers? [LAUGHTER]/nOK. Who says he hasn't proved the case? Why? Exactly. I mean, great, the model fits the data. He had the data first and he made a model to fit it. Big deal. So you would say? Yes, he should be able to make a variety of predictions. That would be a confirmation of a model, at least the beginning of a confirmation of a model is he could make some predictions based on a model./nBut an ex post facto model to explain the data you already have, of course you're going to have one. It might be a little whacky, but you always make a model to explain your data. That's not the hard thing. Now give me some predictions. So, guys, give me some predictions. We write back to Mendel saying we find the author's work to be of interest, it's a provocative and unheard of finding, and it's a fascinating model, but it is just a model. We'd like to see some predictions verified. So what would they be?/nSorry? Color. Oh, show me more traits. OK. Fine. We want to see more traits. In addition to seeing some more traits, and Mendel actually did have more traits in the paper. I'm just simplifying here. Prove this model. What predictions would you make if this model is correct? Yes? Keep crossing them. So tell me what you would do. Please send him instructions here./nOK, so you would like me to cross one of the rounds, an F2 round by a wrinkled. What will happen in the next generation?/nHow do I do that? I don't have DNA sequencing available or anything, so. [LAUGHTER] See what happens. So what might happen? What is this round plant here? What might it be? And what are the probabilities of that?/nOne-third of the time it will big R, big R. Two-thirds of the time it will be big R, little R. If it is big R, big R then the offspring will all be what? Round. If, on the other hand, it is the case that that's big R, little R then the offspring will all be?/nThey won't be all anything. They'll be half round, a 1:1 ratio of round to wrinkled. OK? That's an odd prediction that a third of the time the offspring from such crosses will all be round and two-thirds of the time the offspring will be 50/50 round and wrinkled. You wouldn't normally think of that, right? That's the kind of thing that has to be done. And Mendel, of course, did crosses like that. I simplified here./nThis is really what Mendel did was demonstrated that all sorts of predictions would be satisfied. Another prediction that Mendel could make, oops. Stop, stop, stop, stop. Which should be wrinkled? Oh, my goodness. Oh, wrinkle that pea. OK. Onward. Thank you very much, Claudette. That's good./nSo he made more and more predictions like this. His predictions, for example, let's just take that F1 pea, round over wrinkled here. If you cross this back with wrinkled then it's pretty simply because then always, if this is an F1 as opposed to an F2, you're going to get a 50:50 ratio of round to wrinkled./nMoreover, these rounds, if you cross them back, will still give you a 50:50, etc. That's science. That's the heart of science, is being able to look at data, intuit what the data is trying to tell you, build a model and test a model. All of that is in Mendel. OK? So I know you all know Mendel, but this Mendel really. OK? Now, some definitions. I need to give you, so Section 2, some definitions./nBecause I've been skirting around using some words here. OK? Number one, the word gene. Gene is one of these factors of inheritance controlling a trait./nMendel didn't use the word gene. The word gene came along much later. The variant flavors of a gene, big R and little R, are known as alleles from the Greek word meaning other. These are the alternative forms of a gene./nIt can come in the form big R, little R. I might write big A, little A. I might write plus for normal and M for mutant. There are a lot of different notations geneticists use for that./nThe word phenotype means appearance. The plant was round. The peas were round. That's a phenotype. The individual was 7" 7' tall. That's a phenotype. OK? Those are phenotypes. Genotype means the pair of alleles carried by the individual./nBig R, little R is a genotype. Big R, big R is a genotype. Little R, little R. Those are genotypes. An important difference between genotype and phenotype. Other important words so that we can actually talk to each other. Homozygous or homozygote./nA homozygote is an individual who has a genotype that has two of the same alleles. Two copies of the same allele, the individual is said to be homozygous. And, alternatively, an individual is said to be heterozygous, heterozygote if they have two alternatives./nA couple of other important definitions./nDominant./nA phenotype round is said to be dominant over a phenotype wrinkled if what? If the heterozygote shows that phenotype, the heterozygote between pure breeding strains./nSo phenotype one, pheno one is dominant over phenotype two if the F1 of pure breeding strains shows phenotype one./nSimilarly, we have the word recessive. Now, I'll mention, and you will then proceed to promptly forget, because all of my colleagues forget, dominant and recessive do not refer to alleles. Big R is not dominant. Round is dominant. Big R is an allele. Now, you say who cares? The textbooks get this wrong all the time, it's true. You won't even find the textbooks use this correctly. They will tell you big R is dominant./nWhat if it turned out that big R controlled three different traits? Maybe roundness. An ability to grow with low salt in the soil. An ability to bloom in May. Some of those traits might be recessive. Some of them might be dominant. We know examples of that, where the same allele can control multiple traits, some of which show dominance, some of which show recessiveness. So real card-carrying geneticists try hard to use the word dominant and recessive to refer to phenotypes, not to alleles or genotypes./nNow, since 80% of the facility in the Biology Department don't use the word with that degree of precision, I don't have high hope that you will either. But I'm going to try to say the words dominant and recessive refer to phenotypes. OK? This is a geneticists' kind of hang-up. We all have our shtick, but this one of mine, is that these really do refer to phenotypes. And it's quite important because otherwise you could get quite bollixed up. And I'll come to a case with sickle cell anemia where you won't be able to describe the sickle cell anemia allele as recessive, dominant or co-dominant./nOK? Good. Those are some definitions. They're worth knowing. If we get those definitions right the rest of it is pretty easy. All right./nSo Mendel publishes this paper in 1865. It's accepted. It appears not in Nature but in the proceeding the Royal Academy of Braunau and it's published./nAnd what happens? Nothing. It sinks like a stone. Mendel's paper is totally ignored. Nobody really pays any attention to it. This paper was sent to many people. Charles Darwin has a copy of Mendel's papers in his files. But, in those days, the way printing worked, in order to read a book you had to slit the pages open. Darwin never slit the pages of Mendel's paper, so it's pretty clear he never read the paper, even though it had the answer to much of what he wanted to know about evolution./nNo one really read Mendel's paper because it was so far ahead of its time, it just was pretty strange. It had all these concepts. And, anyway, you could always dismiss it with that kiss of death of biology "it's just the model." Right? You can kill things with "it's just the model." People were just not prepared to deal with Mendel. So Mendel, in fact, poor Mendel, maybe he had a good time, I don't think, instead didn't really do much more on this topic of genetics per se. He became an administrator./nBecame abbot of the monastery and did other things. Worked on meteorology, etc. And we don't really hear from Mendel again. So what really begins to reignite interest in this is the understanding in the late 1800s of chromosomes. Very briefly, cytologists, people studying cells in the microscope. Cytologists are folks who study cells./nThey noticed these very funny little structures in cells. They noticed these structures that when you stain then with a dye would stain very funny. They picked up dye in a certain way. And they noticed that they had this very interesting choreography that when a cell underwent mitosis these funny things would divide down the midline and these little x-shaped structures would go to the two daughter cells like this./nThat is these Xs would become single individual pieces./nAgain, you know about these things. They had no clue what these were. What is the appropriate scientific procedure when you have no clue what something is? You need to give it a name that somewhat covers up the fact that you have no clue what you're talking about because it sounds much better than just saying they are "these funny things." And so they were referred to as chromosomes, meaning literally colored things. [LAUGHTER]/nYou need to understand these sorts of things. OK? So these chromosomes here, these colored things, for lack of any other knowledge of them, that was the property they could be given. Chromosomes. Look it up. They executed this very interesting choreography during mitosis. That is cell division. Oh, boy, is that going to be noisy./nSomeone should shoot it and put it out of its misery. [LAUGHTER] All right. But what they then noticed was the following. And we're going to run just a couple of minutes over. I'm going to keep it short. But they noticed that when organisms made sperm and eggs rather than normal cell division, they noticed that these chromosomes, instead of all of them lining up on the midline, lined up in pairs./nAnd the pairs underwent a series of two divisions. There was a first division which we call meiosis one in which --/n-- one copy of each of these Xs went to each daughter cell./nVery different than mitosis where the Xs would be split down the middle. Then a second division occurred, meiosis two. And in that each of the daughter cells now the X is divided./nAnd they got that./nThis one looked, for all the world, like mitosis. But instead, at the end of the day instead of ending up with four chromosomes, here we end up with only two chromosomes in each gamete, sperm or eggs. And what happened was from this pair, one member of the pair was selected. Now, this is either producing sperm or eggs./nWhen a sperm like that came together with an egg like that and fertilization occurred, you get back to four chromosomes. You all know this. You learned this in high school. But the important point about this was that people said, ha, things lining up in pairs, one copy of each going to the offspring, then a copy from mom and a copy from dad restoring the pair./nSounds just like what that dead monk was talking about. [LAUGHTER] It was just the reason people really didn't think much of Mendel's paper was because it was so abstract. What were these genes? He didn't point to anything. There was nothing concrete. And folks hate that. By contrast they now began to see things and vaguely remembered that this was just like what Mendel's story was about. And three different groups around the world began to redo this work on crosses and all that./nAnd wonderfully in 1900 three groups simultaneously published papers about this. Now, Mendel's Law is rediscovered. Now, the explanation here. How does the cytological observations about meiosis explain Mendel's laws of inheritance of traits? Very simply. All you have to imagine is that big R is being carried on one of these chromosomes./nLittle R on the other one. And then half the offspring had big R, half the offspring had little R. All of Mendel's laws can be implemented by simply assuming that genes and the alleles of those genes live on these chromosomes. So it's beautiful, except for one problem. You may remember from your high schools that Mendel also had another law about more than one trait, pairs of traits./nNot just that we have this segregations of alleles away for one trait. What was his law about two traits? We'll go over this next time. What was his law about two traits, like round and wrinkled and green and yellow? That they would be inherited independently of each other. How would that fit into this model? Different chromosomes. They'd be on different chromosomes./nBut what if I had three traits? Eventually, if I had, now, peas actually have seven pairs of chromosomes. So if I study eight traits in peas, two would have to lie on the same chromosomes. So then the chromosome model would contradict independent inheritance. So either Mendel cannot be right with this other law of independent inheritance that you learned about or the Chromosome Theory cannot be right of these living on these physical molecules and getting distributed that way./nRight? So we have a deep problem because either Mendel, my hero, is wrong or this chromosome model is wrong. And the problem is we don't have enough time to resolve this today, so we're going to have to come back on Wednesday and figure out what happens./nref: www.mit.edu
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Added: March 31, 2009, 7:00 pm
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