1 00:00:00,040 --> 00:00:02,460 The following content is provided under a Creative 2 00:00:02,460 --> 00:00:03,870 Commons license. 3 00:00:03,870 --> 00:00:06,910 Your support will help MIT OpenCourseWare continue to 4 00:00:06,910 --> 00:00:10,560 offer high quality educational resources for free. 5 00:00:10,560 --> 00:00:13,460 To make a donation or view additional materials from 6 00:00:13,460 --> 00:00:19,290 hundreds of MIT courses, visit MIT OpenCourseWare at 7 00:00:19,290 --> 00:00:21,220 ocw.mit.edu. 8 00:00:21,220 --> 00:00:22,550 PROFESSOR: So here's what we did. 9 00:00:22,550 --> 00:00:25,710 We found mutants that effect biochemistry. 10 00:00:25,710 --> 00:00:27,330 That's one way to make the connection between 11 00:00:27,330 --> 00:00:28,830 function and gene. 12 00:00:28,830 --> 00:00:32,920 But if we want to go the other way, we now have to do the 13 00:00:32,920 --> 00:00:35,380 biochemistry of genetics. 14 00:00:35,380 --> 00:00:36,960 What's biochemistry about? 15 00:00:36,960 --> 00:00:39,210 It's purifying things in a test tube. 16 00:00:39,210 --> 00:00:40,810 What's genetics about? 17 00:00:40,810 --> 00:00:42,240 Heredity. 18 00:00:42,240 --> 00:00:43,630 So what do we have to do? 19 00:00:43,630 --> 00:00:48,020 We have to purify heredity in a test tube. 20 00:00:48,020 --> 00:00:48,600 But that's it. 21 00:00:48,600 --> 00:00:49,700 That's what we have to do. 22 00:00:49,700 --> 00:00:51,730 If we're going to make that connection in that direction, 23 00:00:51,730 --> 00:00:55,990 all we need is to take a cell, grind it up, and purify not 24 00:00:55,990 --> 00:00:58,910 the enzyme that digests a sugar, but heredity. 25 00:00:58,910 --> 00:01:02,120 We need to get a pure tube of heredity. 26 00:01:02,120 --> 00:01:04,674 You can imagine that this wasn't an obvious thing to do, 27 00:01:04,674 --> 00:01:07,460 how you get a pure tube of heredity. 28 00:01:07,460 --> 00:01:09,910 The problem is you need an assay. 29 00:01:09,910 --> 00:01:11,470 If you want to find an enzyme that digests a 30 00:01:11,470 --> 00:01:12,710 sugar, you have an acid. 31 00:01:12,710 --> 00:01:14,560 You purify different fraction, you make different fractions 32 00:01:14,560 --> 00:01:17,450 of a cell you see which fraction of the cell is able 33 00:01:17,450 --> 00:01:20,100 to digest the sugar. 34 00:01:20,100 --> 00:01:22,330 But if I make different fractions of the cell, it's 35 00:01:22,330 --> 00:01:24,250 not obvious how I figure out which fraction of the cell has 36 00:01:24,250 --> 00:01:26,780 heredity in it. 37 00:01:26,780 --> 00:01:28,320 And yet, that's exactly what was done, and 38 00:01:28,320 --> 00:01:29,800 that's today's lecture. 39 00:01:29,800 --> 00:01:37,725 So purifying heredity. 40 00:01:42,080 --> 00:01:50,055 The discovery of the transforming principle. 41 00:01:59,880 --> 00:02:02,390 By the transforming principle, I don't mean an 42 00:02:02,390 --> 00:02:04,250 idea, like a principle. 43 00:02:04,250 --> 00:02:05,850 I mean a substance. 44 00:02:05,850 --> 00:02:09,199 This is an old medieval kind of word, a principle. 45 00:02:09,199 --> 00:02:11,400 It's kind of a Harry Potteresque kind of word or 46 00:02:11,400 --> 00:02:14,800 something like that of what is the transforming principle, 47 00:02:14,800 --> 00:02:16,600 the transforming substance. 48 00:02:16,600 --> 00:02:19,050 It's the kind of word alchemists would like to use, 49 00:02:19,050 --> 00:02:21,960 but it was actually what's attached to this, and it was 50 00:02:21,960 --> 00:02:24,610 called at the time the transforming principle, the 51 00:02:24,610 --> 00:02:26,240 transforming substance. 52 00:02:26,240 --> 00:02:31,660 And it really is the work in 1928 of a young scientist F. 53 00:02:31,660 --> 00:02:34,970 Griffiths in London. 54 00:02:34,970 --> 00:02:38,180 Griffiths was particularly interested in studying the 55 00:02:38,180 --> 00:02:39,680 bacteria pneumococcus. 56 00:02:42,410 --> 00:02:45,030 Why was Griffith so interested in studying the bacteria 57 00:02:45,030 --> 00:02:46,500 pneumococcus? 58 00:02:46,500 --> 00:02:50,220 Well, not so long before in 1918, there had been the 59 00:02:50,220 --> 00:02:53,570 terrible Spanish influenza epidemic that had killed 60 00:02:53,570 --> 00:02:57,200 millions of people around the world, the worst flu ever. 61 00:02:57,200 --> 00:03:00,660 Millions of people died by this flew, and one of the 62 00:03:00,660 --> 00:03:03,310 reasons they died when they have the flu was they got 63 00:03:03,310 --> 00:03:06,200 pneumococcal infections. 64 00:03:06,200 --> 00:03:09,140 Griffiths was trying to make a vaccine against pneumococcus, 65 00:03:09,140 --> 00:03:11,870 a pretty good idea trying to make a vaccine against 66 00:03:11,870 --> 00:03:13,130 pneumococcus. 67 00:03:13,130 --> 00:03:17,400 Pneumococcus highly virulent stuff, but particularly if 68 00:03:17,400 --> 00:03:20,220 you're compromised by the influenza. 69 00:03:20,220 --> 00:03:21,510 So what did he do? 70 00:03:21,510 --> 00:03:24,640 Well, Griffiths had a strain of pneumococcus. 71 00:03:24,640 --> 00:03:26,550 He didn't, by the way, infect people with it. 72 00:03:26,550 --> 00:03:29,570 He infected mice with it, that being considered a somewhat 73 00:03:29,570 --> 00:03:31,980 more ethical way to do the experiment. 74 00:03:31,980 --> 00:03:35,610 So he had a strain of pneumococcus that had a 75 00:03:35,610 --> 00:03:37,900 smooth, glistening coat. 76 00:03:37,900 --> 00:03:41,710 When you looked at it, it was smooth and white and 77 00:03:41,710 --> 00:03:43,430 glistened, the colonies that it made 78 00:03:43,430 --> 00:03:44,750 smooth, white, glistened. 79 00:03:44,750 --> 00:03:46,780 And it was virulent. 80 00:03:46,780 --> 00:03:52,350 If you inject it into a mouse, the mouse got pneumococcus 81 00:03:52,350 --> 00:03:54,970 infection, and it died. 82 00:03:54,970 --> 00:03:57,180 It turns out we now know that it has a beautiful 83 00:03:57,180 --> 00:04:00,760 polysaccharide coat around it that provides resistance to 84 00:04:00,760 --> 00:04:03,780 the host's immune system, et cetera. 85 00:04:03,780 --> 00:04:06,610 He also had a strain of pneumococcus 86 00:04:06,610 --> 00:04:09,380 that had a rough coat. 87 00:04:09,380 --> 00:04:10,660 It was not glistening. 88 00:04:10,660 --> 00:04:14,110 It was kind of dull looking, and it was non-virulent. 89 00:04:19,920 --> 00:04:21,570 He injected it into a mouse, mouse lives. 90 00:04:21,570 --> 00:04:23,550 It happens to be the case that we now know that it had a 91 00:04:23,550 --> 00:04:26,190 mutation in a gene that produced that coat, but that 92 00:04:26,190 --> 00:04:27,060 doesn't much matter. 93 00:04:27,060 --> 00:04:29,120 It didn't produce that polysaccharide, and therefore, 94 00:04:29,120 --> 00:04:31,880 was more easily fought off by the immune system. 95 00:04:31,880 --> 00:04:35,130 So Griffiths does the following experiment. 96 00:04:35,130 --> 00:04:47,750 He takes his smooth, virulent bacteria, he injects it into a 97 00:04:47,750 --> 00:04:50,070 mouse, that's a mouse. 98 00:04:50,070 --> 00:04:51,300 And what happens to the mouse? 99 00:04:51,300 --> 00:04:52,620 AUDIENCE: Dies. 100 00:04:52,620 --> 00:04:53,870 PROFESSOR: Dies. 101 00:04:56,510 --> 00:04:57,180 Exactly. 102 00:04:57,180 --> 00:04:59,250 That's a dead mouse. 103 00:04:59,250 --> 00:05:02,220 And one of the easier assays in molecular biology is the 104 00:05:02,220 --> 00:05:04,210 feet up, feet down assay. 105 00:05:04,210 --> 00:05:05,380 All right. 106 00:05:05,380 --> 00:05:06,910 Dead mouse. 107 00:05:06,910 --> 00:05:08,120 Now, what does he do? 108 00:05:08,120 --> 00:05:14,800 He takes the rough, non-virulent, he injects it 109 00:05:14,800 --> 00:05:17,335 into a mouse, and what happens? 110 00:05:20,270 --> 00:05:20,800 Lives. 111 00:05:20,800 --> 00:05:21,460 Right? 112 00:05:21,460 --> 00:05:22,710 Lives. 113 00:05:24,710 --> 00:05:31,790 Then what he does, he takes the smooth, virulent and he 114 00:05:31,790 --> 00:05:33,860 bakes it in an autoclave. 115 00:05:33,860 --> 00:05:36,845 He heat kills this bacteria. 116 00:05:40,070 --> 00:05:47,260 Now, this heat-killed, dead, virulent bacteria when 117 00:05:47,260 --> 00:05:50,770 injected into a mouse, what happens to the mouse? 118 00:05:50,770 --> 00:05:51,650 It's alive. 119 00:05:51,650 --> 00:05:53,130 The mouse is fine. 120 00:05:53,130 --> 00:05:56,000 The bacteria was dead. 121 00:05:56,000 --> 00:06:00,300 And then he does the following truly weird experiment. 122 00:06:00,300 --> 00:06:05,440 He takes the absolutely harmless, rough, non-virulent 123 00:06:05,440 --> 00:06:19,800 bacteria but alive, plus the smooth, virulent bacteria that 124 00:06:19,800 --> 00:06:24,960 has been killed by heat, heat-killed. 125 00:06:24,960 --> 00:06:28,100 The rough stuff is harmless. 126 00:06:28,100 --> 00:06:32,240 The smooth heat-killed stuff, harmless. 127 00:06:32,240 --> 00:06:34,440 Both are harmless. 128 00:06:34,440 --> 00:06:37,690 We've shown the mouse can live with this injected. 129 00:06:37,690 --> 00:06:39,700 It can live with this injected. 130 00:06:39,700 --> 00:06:44,700 He injects it into the mouse, and what happens? 131 00:06:44,700 --> 00:06:45,950 Dead mouse. 132 00:06:51,970 --> 00:06:54,350 Very surprising. 133 00:06:54,350 --> 00:06:57,320 Not only that, when he takes the blood of the dead mouse, 134 00:06:57,320 --> 00:07:00,130 he can culture out of it. 135 00:07:00,130 --> 00:07:04,370 He can culture on a Petri plate from this dead mouse 136 00:07:04,370 --> 00:07:08,760 live, smooth, virulent bacteria. 137 00:07:13,090 --> 00:07:15,970 How did that happen? 138 00:07:15,970 --> 00:07:19,420 Somehow, we didn't have any live, smooth bacteria. 139 00:07:19,420 --> 00:07:20,575 We had dead, smooth bacteria. 140 00:07:20,575 --> 00:07:22,750 We had live non-virulent bacteria. 141 00:07:22,750 --> 00:07:29,680 Somehow, the dead stuff transformed the live harmless 142 00:07:29,680 --> 00:07:32,120 stuff into virulence. 143 00:07:32,120 --> 00:07:34,030 It transformed it. 144 00:07:34,030 --> 00:07:38,030 The substance, the unknown substance that transformed it 145 00:07:38,030 --> 00:07:40,980 was referred to as the transforming principle or 146 00:07:40,980 --> 00:07:43,000 transforming stuff. 147 00:07:43,000 --> 00:07:47,840 And now we have biochemistry because we have an assay. 148 00:07:47,840 --> 00:07:52,660 We could take the dead, virulent bacteria and break it 149 00:07:52,660 --> 00:07:55,120 up into fractions and see which substance, which 150 00:07:55,120 --> 00:07:56,050 fraction, is it a protein? 151 00:07:56,050 --> 00:07:56,880 Is it a carbohydrate? 152 00:07:56,880 --> 00:07:57,840 Is it nucleic acid? 153 00:07:57,840 --> 00:08:02,090 Is it something, is the transforming substance and 154 00:08:02,090 --> 00:08:05,960 purify heredity because we have seen the transfer of 155 00:08:05,960 --> 00:08:08,950 heredity to the harmless bacteria. 156 00:08:08,950 --> 00:08:09,590 There is an assay. 157 00:08:09,590 --> 00:08:12,530 The minute there's an assay, you could do biochemistry. 158 00:08:12,530 --> 00:08:17,060 Now, the problem was the assay was painfully slow. 159 00:08:17,060 --> 00:08:21,370 You had to grind up the dead bacteria, you had to 160 00:08:21,370 --> 00:08:24,300 demonstrate by mixing it together, putting it into a 161 00:08:24,300 --> 00:08:28,240 mouse, waiting months, and then you had to get a further 162 00:08:28,240 --> 00:08:29,020 sub fraction. 163 00:08:29,020 --> 00:08:31,000 It was just painfully slow. 164 00:08:31,000 --> 00:08:32,860 All this mouse stuff and all that. 165 00:08:32,860 --> 00:08:37,070 And Griffiths didn't make much progress through the 1930s, 166 00:08:37,070 --> 00:08:38,820 but he kept going at it. 167 00:08:38,820 --> 00:08:43,360 And I suspect might have gotten there except for the 168 00:08:43,360 --> 00:08:48,850 fact that in 1941, his lab was hit by a German bomb during a 169 00:08:48,850 --> 00:08:51,360 blitz, and he died. 170 00:08:51,360 --> 00:08:57,440 And so Griffiths never saw the result of this, but he did 171 00:08:57,440 --> 00:08:59,620 purify fractions and all that. 172 00:08:59,620 --> 00:09:02,410 But thank goodness others picked it up. 173 00:09:02,410 --> 00:09:04,180 He was again, Griffiths was a great guy. 174 00:09:04,180 --> 00:09:08,250 He worked in World War I, worked during World War II on 175 00:09:08,250 --> 00:09:11,370 important public health problems, and really lays this 176 00:09:11,370 --> 00:09:14,660 foundation, but never really purified what the final 177 00:09:14,660 --> 00:09:18,240 substance was because he died in a bombing. 178 00:09:18,240 --> 00:09:24,060 But then you get in 1943, during World War II, folks 179 00:09:24,060 --> 00:09:27,930 working in New York City at Rockefeller University, then 180 00:09:27,930 --> 00:09:36,040 Rockefeller Institute, Avery, McCarty, and MacLeod continue 181 00:09:36,040 --> 00:09:41,620 this work, and they do it without the mouse. 182 00:09:41,620 --> 00:09:45,570 Because what they do is they grind up the virulent stuff 183 00:09:45,570 --> 00:09:49,180 and they sprinkle it on the living stuff, the living 184 00:09:49,180 --> 00:09:52,290 bacteria and played it out, and just looked for the 185 00:09:52,290 --> 00:09:56,370 appearance of colonies that are transformed. 186 00:09:56,370 --> 00:09:57,980 Skip the mouse. 187 00:09:57,980 --> 00:10:00,790 Skipping the mouse makes it a lot easier, because you can do 188 00:10:00,790 --> 00:10:02,080 those experiments pretty quickly. 189 00:10:02,080 --> 00:10:03,570 Bacteria grow quickly. 190 00:10:03,570 --> 00:10:05,090 Same basic idea. 191 00:10:05,090 --> 00:10:10,830 Purify the stuff from the smooth dead stuff, grind it 192 00:10:10,830 --> 00:10:13,740 up, put it in different fractions, apply them, and 193 00:10:13,740 --> 00:10:15,900 then sprinkle them on a plate, and look for 194 00:10:15,900 --> 00:10:18,120 the occasional colony. 195 00:10:18,120 --> 00:10:20,300 And now you're just going to look for a fraction of the 196 00:10:20,300 --> 00:10:22,970 material that has the capability to produce some 197 00:10:22,970 --> 00:10:25,460 smooth colonies. 198 00:10:25,460 --> 00:10:27,270 Well, they did that, and they purified it, and the purified 199 00:10:27,270 --> 00:10:30,060 it, and they purified it, and they purified it, and they 200 00:10:30,060 --> 00:10:35,230 eventually found that the particular type of molecule 201 00:10:35,230 --> 00:10:40,280 that they purified appeared to be DNA. 202 00:10:40,280 --> 00:10:52,660 But when they purified fractions containing DNA, 203 00:10:52,660 --> 00:10:57,620 these fractions had the ability to transform. 204 00:11:03,360 --> 00:11:04,610 Wow. 205 00:11:08,080 --> 00:11:11,100 You might immediately say, that's the transforming 206 00:11:11,100 --> 00:11:12,070 principle, DNA. 207 00:11:12,070 --> 00:11:15,130 That's the transforming substance. 208 00:11:15,130 --> 00:11:16,380 What do you think the reaction was? 209 00:11:19,910 --> 00:11:21,220 Skepticism. 210 00:11:21,220 --> 00:11:23,590 First, it should be noted it's 1943. 211 00:11:23,590 --> 00:11:26,210 People were busy at the time, right? 212 00:11:26,210 --> 00:11:28,040 We're in the middle World War II. 213 00:11:28,040 --> 00:11:31,280 It wasn't exactly top on people's minds, but there was 214 00:11:31,280 --> 00:11:34,430 enormous skepticism scientifically of those people 215 00:11:34,430 --> 00:11:36,420 who did follow the work. 216 00:11:36,420 --> 00:11:38,950 Why? 217 00:11:38,950 --> 00:11:45,020 Because the one thing they knew was that DNA was truly a 218 00:11:45,020 --> 00:11:46,770 boring molecule. 219 00:11:46,770 --> 00:11:50,330 It was understood by all smart people that DNA was an 220 00:11:50,330 --> 00:11:54,200 incredibly boring structural molecule that had none of the 221 00:11:54,200 --> 00:11:57,840 fascinating diversity and richness of proteins. 222 00:11:57,840 --> 00:12:00,660 Proteins could do zillions of different things. 223 00:12:00,660 --> 00:12:03,730 DNA, you know, it's just scaffolding. 224 00:12:03,730 --> 00:12:04,930 Why? 225 00:12:04,930 --> 00:12:07,090 What is the structure of DNA? 226 00:12:07,090 --> 00:12:11,310 So let's turn to the structure of DNA to see why it is that 227 00:12:11,310 --> 00:12:14,570 people were not impressed. 228 00:12:14,570 --> 00:12:17,210 Of course, when people are not impressed, you purified 229 00:12:17,210 --> 00:12:20,580 something and you show it transforms, what do you say to 230 00:12:20,580 --> 00:12:24,000 Avery, McCarty, and MacLeod? 231 00:12:24,000 --> 00:12:26,610 How do you express your skepticism? 232 00:12:26,610 --> 00:12:28,360 You say, it's very good. 233 00:12:28,360 --> 00:12:31,220 You've purified this and it contains DNA, but is it 234 00:12:31,220 --> 00:12:35,570 absolutely totally 100% pure or is it possible that you've 235 00:12:35,570 --> 00:12:39,550 carried along in the fraction that you have purified some 236 00:12:39,550 --> 00:12:42,610 other trace quantity of a highly potent protein that is 237 00:12:42,610 --> 00:12:45,290 really causing heredity? 238 00:12:45,290 --> 00:12:48,200 And of course, that's the problem is you can never prove 239 00:12:48,200 --> 00:12:50,950 that there's not a teeny smidgen of something in there. 240 00:12:50,950 --> 00:12:53,280 You can only show how pure it is, but you can never rule 241 00:12:53,280 --> 00:12:54,060 something out. 242 00:12:54,060 --> 00:12:56,490 So when people want to sort of dis your biochemistry 243 00:12:56,490 --> 00:12:59,480 experiments, it's always easy to say, it was probably 244 00:12:59,480 --> 00:13:02,290 something else in there too you just don't know about it. 245 00:13:02,290 --> 00:13:04,230 And that was what the answer was to them. 246 00:13:04,230 --> 00:13:06,300 But let's look at the structure of DNA. 247 00:13:06,300 --> 00:13:08,580 So DNA has three important components 248 00:13:08,580 --> 00:13:10,050 which we need to learn. 249 00:13:10,050 --> 00:13:15,095 A, it has a sugar called 2 prime deoxyribose. 250 00:13:19,800 --> 00:13:24,940 So ribose is a 5-carbon sugar. 251 00:13:24,940 --> 00:13:27,590 A five-part carbon sugar is a pentose. 252 00:13:27,590 --> 00:13:31,770 So this is a sugar, in fact a pentose, pentose of course 253 00:13:31,770 --> 00:13:36,600 five, pentose meaning it's a 5-carbon sugar, but it lacks a 254 00:13:36,600 --> 00:13:37,870 hydroxyl group. 255 00:13:37,870 --> 00:13:40,490 So it's just slightly different from 256 00:13:40,490 --> 00:13:41,690 the 5-carbon sugar. 257 00:13:41,690 --> 00:13:46,485 And we draw it in this configuration where there is 258 00:13:46,485 --> 00:13:52,620 the 1 prime carbon here, the 2 prime carbon here, the 3 prime 259 00:13:52,620 --> 00:13:57,660 carbon here, the 4 prime carbon here, the 5 prime 260 00:13:57,660 --> 00:13:58,940 carbon here. 261 00:13:58,940 --> 00:14:00,820 That's very important to know. 262 00:14:00,820 --> 00:14:02,550 We've got an oxygen up here. 263 00:14:02,550 --> 00:14:10,200 Here, we have an OH and an H. Here, we have an H and an OH. 264 00:14:10,200 --> 00:14:20,990 Here we have H. Here we have our OH, H, H. 265 00:14:20,990 --> 00:14:25,040 But here we should have a hydroxyl off every 266 00:14:25,040 --> 00:14:29,110 carbon, and we don't. 267 00:14:29,110 --> 00:14:35,280 Only here are deoxy. 268 00:14:35,280 --> 00:14:37,030 That's the only difference from this being a perfectly 269 00:14:37,030 --> 00:14:41,440 normal ribose, deoxy at the 2 prime position. 270 00:14:41,440 --> 00:14:43,390 Big deal. 271 00:14:43,390 --> 00:14:51,260 Now, the next component of DNA that you need to know about 272 00:14:51,260 --> 00:14:55,890 are these nitrogenous bases. 273 00:14:55,890 --> 00:15:08,730 So hanging off our ribose is a base. 274 00:15:18,920 --> 00:15:24,940 This base has carbons, oxygens, 275 00:15:24,940 --> 00:15:26,920 hydrogens, and nitrogens. 276 00:15:26,920 --> 00:15:42,830 And they come in four flavors, adenine, A, guanine, G, 277 00:15:42,830 --> 00:15:54,880 thymine, T, cytosine, C. A, T, C and G, and we'll look at 278 00:15:54,880 --> 00:15:57,700 their structure in just a moment. 279 00:15:57,700 --> 00:16:03,690 Then the next important part if we look at this 280 00:16:03,690 --> 00:16:18,450 conceptually, is that hanging off here, we have a 281 00:16:18,450 --> 00:16:19,700 triphosphate. 282 00:16:35,080 --> 00:16:38,290 We have a triphosphate. 283 00:16:38,290 --> 00:16:44,315 So this is the monomer for making DNA, triphosphate. 284 00:16:47,240 --> 00:16:52,110 We have a sugar, the sugar in exactly the same place off the 285 00:16:52,110 --> 00:16:57,900 1 prime carbon there has a base, off the 5 prime carbon, 286 00:16:57,900 --> 00:17:00,090 we have a triphosphate. 287 00:17:00,090 --> 00:17:04,240 What's the triphosphate good for? 288 00:17:04,240 --> 00:17:04,859 Energy. 289 00:17:04,859 --> 00:17:08,200 We're going to do a polymerization, and that's 290 00:17:08,200 --> 00:17:13,010 going to contribute the energy for the polymerization, and 291 00:17:13,010 --> 00:17:14,700 that's pretty much it. 292 00:17:14,700 --> 00:17:16,089 That's the way to think about DNA. 293 00:17:16,089 --> 00:17:22,690 So when we do our polymerization now, we 294 00:17:22,690 --> 00:17:44,706 polymerize and we get base, CH2, phosphate. 295 00:17:50,070 --> 00:17:55,050 And then coming down this way attached here, we have our 296 00:17:55,050 --> 00:18:12,890 phosphate, and that attaches to the 5 prime carbon here, 297 00:18:12,890 --> 00:18:14,840 and onward that way. 298 00:18:14,840 --> 00:18:28,170 So notice that our polymer goes from a 5-prime carbon 299 00:18:28,170 --> 00:18:35,710 here, 3-prime carbon here, 5-prime carbon here, 3-prime 300 00:18:35,710 --> 00:18:37,390 carbon there. 301 00:18:37,390 --> 00:18:41,530 And we go sugar, phosphate, sugar, phosphate, sugar, 302 00:18:41,530 --> 00:18:45,240 phosphate, sugar, phosphate, 5-prime attachment, 3-prime 303 00:18:45,240 --> 00:18:47,430 attachment, 5-prime attachment, 3-prime 304 00:18:47,430 --> 00:18:48,370 attachment. 305 00:18:48,370 --> 00:18:50,680 That's DNA. 306 00:18:50,680 --> 00:18:52,630 Pretty boring. 307 00:18:52,630 --> 00:18:57,060 The same sugar, same phosphates strung together, 308 00:18:57,060 --> 00:18:58,850 totally boring. 309 00:18:58,850 --> 00:19:02,270 The only difference are these bases. 310 00:19:02,270 --> 00:19:04,860 And there's only four of them, and they're not very 311 00:19:04,860 --> 00:19:06,582 impressive. 312 00:19:06,582 --> 00:19:10,460 They're pretty boring, these bases. 313 00:19:10,460 --> 00:19:12,920 There are purines. 314 00:19:12,920 --> 00:19:15,900 The A and G are purines, and their ring 315 00:19:15,900 --> 00:19:18,750 structure looks like this. 316 00:19:18,750 --> 00:19:24,590 This is six-membered ring and there's a five-membered ring. 317 00:19:24,590 --> 00:19:31,870 There are pyrimidines, T and C, and they just have a 318 00:19:31,870 --> 00:19:33,120 six-membered ring. 319 00:19:39,600 --> 00:19:42,890 They've got carbons, and oxygens, and nitrogens, and 320 00:19:42,890 --> 00:19:46,510 hydrogens, and they don't differ 321 00:19:46,510 --> 00:19:48,630 really in their charges. 322 00:19:48,630 --> 00:19:51,670 By compared to the amino acids, positive charges, 323 00:19:51,670 --> 00:19:53,820 negative charges, hydrophobic groups, 324 00:19:53,820 --> 00:19:55,660 sulfurs that are reactive. 325 00:19:55,660 --> 00:19:57,230 Amino acids, that's impressive. 326 00:19:57,230 --> 00:20:00,030 Those 20 different side chains have wildly different chemical 327 00:20:00,030 --> 00:20:00,820 properties. 328 00:20:00,820 --> 00:20:04,510 These form measly bases, have essentially, the same chemical 329 00:20:04,510 --> 00:20:05,300 properties. 330 00:20:05,300 --> 00:20:07,240 There's nothing very different about their chemical 331 00:20:07,240 --> 00:20:10,410 properties and therefore, all smart right-thinking people 332 00:20:10,410 --> 00:20:12,220 recognize the DNA could not be a 333 00:20:12,220 --> 00:20:14,800 particularly interesting molecule. 334 00:20:14,800 --> 00:20:17,020 It had to be largely a structural 335 00:20:17,020 --> 00:20:18,850 molecule of some sort. 336 00:20:18,850 --> 00:20:23,400 So when Avery, McCarty, and MacLeod tell us ah, the 337 00:20:23,400 --> 00:20:26,695 transforming principle of DNA, nobody's impressed. 338 00:20:31,020 --> 00:20:33,500 But of course, it's World War II. 339 00:20:33,500 --> 00:20:35,020 People are busy. 340 00:20:35,020 --> 00:20:36,670 Lot of things going on. 341 00:20:36,670 --> 00:20:41,810 And not that long afterwards, not that long afterwards, 342 00:20:41,810 --> 00:20:44,400 another really important experiment gets done in the 343 00:20:44,400 --> 00:20:56,515 early 1950s, the Hershey-Chase experiment. 344 00:21:03,560 --> 00:21:05,260 Hershey is not the candy bar. 345 00:21:05,260 --> 00:21:09,500 It is Alfred Hershey and Martha Chase. 346 00:21:09,500 --> 00:21:14,680 Martha Chase and Alfred Hershey do a cool experiment. 347 00:21:14,680 --> 00:21:19,780 People were studying something else at the time. 348 00:21:19,780 --> 00:21:26,060 They were studying the viruses that infect bacteria, 349 00:21:26,060 --> 00:21:28,470 bacterial viruses. 350 00:21:28,470 --> 00:21:35,670 So it turns out just like you may get a viral infection, E. 351 00:21:35,670 --> 00:21:38,540 coli gets viral infections too. 352 00:21:38,540 --> 00:21:41,440 It usually dies of them or at least often dies of viral 353 00:21:41,440 --> 00:21:43,830 infection, not necessarily usually, I take that back, 354 00:21:43,830 --> 00:21:47,300 sometimes dies of viral infections. 355 00:21:47,300 --> 00:21:54,120 So that is a virus, actually, greatly magnified, glommed on 356 00:21:54,120 --> 00:21:57,710 to E. coli, virus E. coli. 357 00:22:00,310 --> 00:22:06,560 What happens is, if you mix the virus with E. coli, it 358 00:22:06,560 --> 00:22:10,850 gloms on, and then if you wait a little while giving it a 359 00:22:10,850 --> 00:22:14,880 happy medium to grow in, the E. coli some time later, half 360 00:22:14,880 --> 00:22:20,760 an hour later maybe, bursts open, dead, and spews out 361 00:22:20,760 --> 00:22:24,140 zillions of viral particles which could go on 362 00:22:24,140 --> 00:22:26,080 to infect new cells. 363 00:22:26,080 --> 00:22:28,180 How does it do that? 364 00:22:28,180 --> 00:22:32,470 How does it instruct E. coli to make viral particles? 365 00:22:32,470 --> 00:22:34,070 It must be bringing information. 366 00:22:34,070 --> 00:22:36,080 It's having progeny. 367 00:22:36,080 --> 00:22:39,100 It is passing on heredity too. 368 00:22:39,100 --> 00:22:41,940 It has some transforming information. 369 00:22:41,940 --> 00:22:47,310 Where is the transforming principle in the little virus? 370 00:22:47,310 --> 00:22:50,560 It gloms on to the cell somehow gives something into 371 00:22:50,560 --> 00:22:56,210 the cell, and poof, 20 minutes later, half an hour later, 372 00:22:56,210 --> 00:22:58,060 lots of viruses. 373 00:22:58,060 --> 00:22:59,360 Where's the information carried? 374 00:22:59,360 --> 00:23:02,030 Now, this was a much simpler system. 375 00:23:02,030 --> 00:23:05,780 This system, you're asking what's in the bacterial virus. 376 00:23:05,780 --> 00:23:08,170 There's not a lot in a bacterial virus. 377 00:23:08,170 --> 00:23:10,750 It's not like a cell that have zillions of things. 378 00:23:10,750 --> 00:23:14,110 The bacterial virus is a pretty simple particle. 379 00:23:14,110 --> 00:23:32,890 The bacteria virus consists of protein coat, proteins are on 380 00:23:32,890 --> 00:23:38,305 the outside, DNA on the inside. 381 00:23:41,370 --> 00:23:42,490 That's it. 382 00:23:42,490 --> 00:23:44,070 You don't have a lot to work with, a limited number of 383 00:23:44,070 --> 00:23:47,390 proteins, DNA in the middle. 384 00:23:47,390 --> 00:23:50,890 These things just as an aside were thought to kind of like 385 00:23:50,890 --> 00:23:53,030 eat bacteria. 386 00:23:53,030 --> 00:23:56,040 Because they were thought to eat bacteria in a way by at 387 00:23:56,040 --> 00:24:00,470 least the early things, they're called bacteriophage, 388 00:24:00,470 --> 00:24:01,720 bacteriophage. 389 00:24:04,370 --> 00:24:06,390 The word phage means to eat. 390 00:24:06,390 --> 00:24:11,690 So you may hear me talking about bacteriophage, meaning 391 00:24:11,690 --> 00:24:13,520 eaters of bacteria. 392 00:24:13,520 --> 00:24:16,620 Indeed, actually there was some nutty ideas in the 1920s 393 00:24:16,620 --> 00:24:19,860 and 1930s when bacteriophage were first discovered that the 394 00:24:19,860 --> 00:24:22,710 way to cure a bacterial infection was to drink a lot 395 00:24:22,710 --> 00:24:23,960 of bacteriophage. 396 00:24:25,920 --> 00:24:27,660 They would kill the bacteria. 397 00:24:27,660 --> 00:24:28,190 It's a thought. 398 00:24:28,190 --> 00:24:29,570 People actually tried these things. 399 00:24:29,570 --> 00:24:33,830 Anyway, it turns out not to be such a good idea. 400 00:24:33,830 --> 00:24:38,970 So Hershey and Chase decided we're going to figure out 401 00:24:38,970 --> 00:24:39,690 which is it? 402 00:24:39,690 --> 00:24:44,650 Is it the DNA or is it the protein? 403 00:24:48,010 --> 00:24:50,560 How do you find out? 404 00:24:50,560 --> 00:24:50,820 Yeah? 405 00:24:50,820 --> 00:24:52,580 AUDIENCE: [INAUDIBLE]. 406 00:24:52,580 --> 00:24:55,480 PROFESSOR: Put in only protein, and see what happens. 407 00:24:55,480 --> 00:25:00,930 So take the bacteriophage, purify it from 408 00:25:00,930 --> 00:25:03,500 protein verses DNA. 409 00:25:03,500 --> 00:25:08,730 I've got a pure component of the protein, I sprinkle it on, 410 00:25:08,730 --> 00:25:10,800 nothing happens. 411 00:25:10,800 --> 00:25:17,080 I take the DNA, I sprinkle it on, nothing happens. 412 00:25:17,080 --> 00:25:18,920 Neither works. 413 00:25:18,920 --> 00:25:20,170 Why is that? 414 00:25:22,278 --> 00:25:25,659 AUDIENCE: [INAUDIBLE]. 415 00:25:25,659 --> 00:25:29,370 PROFESSOR: The shape, those little feet in the shape were 416 00:25:29,370 --> 00:25:32,060 critical for the pathogenicity. 417 00:25:32,060 --> 00:25:35,840 So when we grind up the virus, it doesn't work anymore. 418 00:25:35,840 --> 00:25:36,800 It's a great idea. 419 00:25:36,800 --> 00:25:39,740 If it worked, bingo, we'd have it, and that should be the 420 00:25:39,740 --> 00:25:42,400 first experiment we do because it's so easy. 421 00:25:42,400 --> 00:25:44,430 But it turned out not to work. 422 00:25:44,430 --> 00:25:44,830 Yes? 423 00:25:44,830 --> 00:25:47,315 AUDIENCE: [INAUDIBLE]. 424 00:25:47,315 --> 00:25:50,680 PROFESSOR: Put a chemical marker on the protein, put a 425 00:25:50,680 --> 00:25:53,490 chemical marker on the DNA, and see which one 426 00:25:53,490 --> 00:25:57,000 goes into the cell. 427 00:25:57,000 --> 00:25:58,250 What chemical marker? 428 00:26:00,700 --> 00:26:05,370 How are we going to attach a chemical marker to the protein 429 00:26:05,370 --> 00:26:06,330 without messing it up? 430 00:26:06,330 --> 00:26:07,580 We can't mess up the protein, right? 431 00:26:07,580 --> 00:26:08,660 It still got to function. 432 00:26:08,660 --> 00:26:10,520 How do we get a chemical marker on it? 433 00:26:10,520 --> 00:26:12,176 AUDIENCE: [INAUDIBLE]. 434 00:26:12,176 --> 00:26:12,845 PROFESSOR: Sorry? 435 00:26:12,845 --> 00:26:14,095 AUDIENCE: [INAUDIBLE]. 436 00:26:17,348 --> 00:26:19,150 PROFESSOR: So what chemical do you want me to put in? 437 00:26:22,690 --> 00:26:25,910 Well, how am I going to tell whether, DNA's got phosphorus. 438 00:26:25,910 --> 00:26:28,654 How am I going to follow the phosphorus? 439 00:26:28,654 --> 00:26:30,030 AUDIENCE: Radioactive tag. 440 00:26:30,030 --> 00:26:32,330 PROFESSOR: Radioactive tag. 441 00:26:32,330 --> 00:26:33,250 Bingo. 442 00:26:33,250 --> 00:26:37,650 What if I used radioactive tags, and I made a 443 00:26:37,650 --> 00:26:40,980 radioactively-labeled virus. 444 00:26:40,980 --> 00:26:43,720 How can I radioactively label the DNA? 445 00:26:43,720 --> 00:26:46,100 AUDIENCE: [INAUDIBLE]. 446 00:26:46,100 --> 00:26:46,850 PROFESSOR: Sorry? 447 00:26:46,850 --> 00:26:48,586 AUDIENCE: [INAUDIBLE] 448 00:26:48,586 --> 00:26:51,260 PROFESSOR: A radioactive base. 449 00:26:51,260 --> 00:26:52,330 I could do that. 450 00:26:52,330 --> 00:26:53,960 What else could I do? 451 00:26:53,960 --> 00:26:54,360 Yup? 452 00:26:54,360 --> 00:26:55,080 AUDIENCE: Phosphorus. 453 00:26:55,080 --> 00:26:55,930 PROFESSOR: Phosphorus. 454 00:26:55,930 --> 00:26:58,470 Phosphorus has the nice property that phosphorus is in 455 00:26:58,470 --> 00:27:00,750 my DNA, but it's not an proteins. 456 00:27:00,750 --> 00:27:01,630 So what do I use? 457 00:27:01,630 --> 00:27:04,110 Phosphorus-32, P-32. 458 00:27:04,110 --> 00:27:07,820 So I use P-32, and how do I manage to chemically create a 459 00:27:07,820 --> 00:27:10,040 virus that has P-32 in it? 460 00:27:10,040 --> 00:27:11,330 AUDIENCE: [INAUDIBLE]. 461 00:27:11,330 --> 00:27:14,060 PROFESSOR: Just throw it in the solution with P-32, and 462 00:27:14,060 --> 00:27:16,410 the virus will take care that itself, right? 463 00:27:16,410 --> 00:27:21,610 So simply grow virus for a while in the presence of P-32. 464 00:27:21,610 --> 00:27:23,730 Let's do that. 465 00:27:23,730 --> 00:27:29,290 So grow virus in a test tube with bacteria. 466 00:27:29,290 --> 00:27:30,740 Here's my bacteria. 467 00:27:30,740 --> 00:27:35,220 Here's my virus I've put in there, and let me put in P-32, 468 00:27:35,220 --> 00:27:41,640 and what I'll get is P-32-labeled labeled virus. 469 00:27:41,640 --> 00:27:42,890 How do I label my protein? 470 00:27:48,440 --> 00:27:49,880 Someone said it already. 471 00:27:49,880 --> 00:27:51,650 What elements can we find that's in 472 00:27:51,650 --> 00:27:53,462 proteins but not in DNA? 473 00:27:53,462 --> 00:27:54,938 AUDIENCE: [INAUDIBLE] 474 00:27:54,938 --> 00:27:55,705 PROFESSOR: Sorry? 475 00:27:55,705 --> 00:27:56,385 AUDIENCE: Sulfur. 476 00:27:56,385 --> 00:27:57,870 PROFESSOR: Sulfur. 477 00:27:57,870 --> 00:28:00,380 Sulfur. 478 00:28:00,380 --> 00:28:05,370 S-35 is a radioactive isotope of sulfur, and if I grow it, I 479 00:28:05,370 --> 00:28:10,870 can S-35 label the proteins in my virus. 480 00:28:16,420 --> 00:28:17,920 Nice. 481 00:28:17,920 --> 00:28:20,420 This radioactive labeling trick is very cool. 482 00:28:20,420 --> 00:28:25,340 So I take it, I take some P-32-labeled virus where these 483 00:28:25,340 --> 00:28:26,910 P-32 was only in the DNA. 484 00:28:26,910 --> 00:28:30,110 I got some S-35-labeled label virus where the 485 00:28:30,110 --> 00:28:31,890 S-35 is in the protein. 486 00:28:31,890 --> 00:28:38,160 I could mix them together, now do my experiment, wait 20 487 00:28:38,160 --> 00:28:43,220 minutes and, or even wait last 10, 15 minutes, and see which 488 00:28:43,220 --> 00:28:47,070 element has gone into the cell. 489 00:28:47,070 --> 00:28:48,320 How do I do that? 490 00:28:50,510 --> 00:28:55,620 See I've got my cells here, and I've got the viruses 491 00:28:55,620 --> 00:28:59,350 attached to them, and they've injected something in here. 492 00:28:59,350 --> 00:29:02,010 They've either injected a protein or 493 00:29:02,010 --> 00:29:05,560 they've injected DNA. 494 00:29:05,560 --> 00:29:07,940 What was injected? 495 00:29:07,940 --> 00:29:12,290 I need to carefully go in there and remove the virus and 496 00:29:12,290 --> 00:29:13,640 look at just what's in the cell. 497 00:29:13,640 --> 00:29:16,880 I have to now separate the virus glommed onto the outside 498 00:29:16,880 --> 00:29:19,580 of the cell from the cell. 499 00:29:19,580 --> 00:29:22,070 So do I use micro manipulator tweezers to 500 00:29:22,070 --> 00:29:23,522 pull off the virus? 501 00:29:23,522 --> 00:29:25,250 AUDIENCE: [INAUDIBLE]. 502 00:29:25,250 --> 00:29:26,930 PROFESSOR: Well, if I denature, I might 503 00:29:26,930 --> 00:29:29,600 crack open the cell. 504 00:29:29,600 --> 00:29:30,110 Centrifuge it. 505 00:29:30,110 --> 00:29:34,080 If I centrifuge it, the whole thing will spin down. 506 00:29:34,080 --> 00:29:38,040 I need to kind of knock the viruses off the cell, 507 00:29:38,040 --> 00:29:38,550 physically. 508 00:29:38,550 --> 00:29:41,520 I just got to agitate it so I get them off the cell. 509 00:29:41,520 --> 00:29:46,450 With enough kind of hydrodynamic agitation, the 510 00:29:46,450 --> 00:29:48,980 viruses fall off. 511 00:29:48,980 --> 00:29:54,310 So a device was created that was able to just perfectly 512 00:29:54,310 --> 00:29:58,140 knock the viruses off. 513 00:29:58,140 --> 00:30:02,240 It's referred to as the Waring kitchen blender. 514 00:30:02,240 --> 00:30:05,620 It turns on your kitchen blender is perfect for this. 515 00:30:05,620 --> 00:30:08,780 Take the viruses, add it to the bacteria, sit for a little 516 00:30:08,780 --> 00:30:12,470 bit, put it in your kitchen blender, press puree. 517 00:30:12,470 --> 00:30:17,240 And let's say on puree setting, the viruses fall off, 518 00:30:17,240 --> 00:30:20,770 and now you can spin it in a centrifuge, the bacteria are 519 00:30:20,770 --> 00:30:22,370 denser, they come down. 520 00:30:22,370 --> 00:30:25,600 The viruses are lighter, they stay in the supernatant, and 521 00:30:25,600 --> 00:30:29,760 you can take the supernatant and the pellet at the bottom 522 00:30:29,760 --> 00:30:32,920 over your radioactivity counter and see which one is 523 00:30:32,920 --> 00:30:34,220 in the bacteria. 524 00:30:34,220 --> 00:30:36,600 These were referred to as the famous Waring blender 525 00:30:36,600 --> 00:30:37,330 experiments. 526 00:30:37,330 --> 00:30:39,310 They really are, actually. 527 00:30:39,310 --> 00:30:44,320 So you put this in the Waring blender, you knock off the 528 00:30:44,320 --> 00:30:54,710 viruses, you spin it down, and what happens is after you've 529 00:30:54,710 --> 00:30:58,150 done it, there's a pellet here of the bacteria that are spun 530 00:30:58,150 --> 00:31:00,200 down in the centrifuge. 531 00:31:00,200 --> 00:31:03,440 The virus particles are still up here. 532 00:31:03,440 --> 00:31:07,880 We take this pellet over to our counter, and which element 533 00:31:07,880 --> 00:31:12,165 do we find in great abundance, S-35 or P-32? 534 00:31:12,165 --> 00:31:12,975 AUDIENCE: P-32. 535 00:31:12,975 --> 00:31:17,740 PROFESSOR: P-32. 536 00:31:17,740 --> 00:31:19,940 The DNA is what's going in. 537 00:31:19,940 --> 00:31:21,500 Bingo. 538 00:31:21,500 --> 00:31:24,510 Nice experiment. 539 00:31:24,510 --> 00:31:27,190 Now if you were churlish, couldn't you say, yeah, look 540 00:31:27,190 --> 00:31:29,600 it's mostly the DNA, but there's a little smidgen of 541 00:31:29,600 --> 00:31:31,480 protein maybe, that came along too. 542 00:31:31,480 --> 00:31:35,770 Do you think they found absolutely zero S-35 in there? 543 00:31:35,770 --> 00:31:37,910 No, because they don't perfectly knock the virus off. 544 00:31:37,910 --> 00:31:39,160 Some of it kind of sticks. 545 00:31:39,160 --> 00:31:41,180 There's 1% S-35. 546 00:31:41,180 --> 00:31:43,770 And if you're being really churlish about this you would 547 00:31:43,770 --> 00:31:45,490 say I still don't believe you. 548 00:31:45,490 --> 00:31:47,400 But now you have it from two different directions. 549 00:31:47,400 --> 00:31:50,600 You have it from the pneumococcus, this bacteria 550 00:31:50,600 --> 00:31:52,940 experiment from Avery, McCarty, and MacLeod Hershey 551 00:31:52,940 --> 00:31:54,930 and Chase coming from two different systems. 552 00:31:54,930 --> 00:31:56,710 They're giving you the same answer. 553 00:31:56,710 --> 00:31:57,620 It's pretty clear. 554 00:31:57,620 --> 00:31:58,250 It's in the air. 555 00:31:58,250 --> 00:32:00,810 People know DNA is the stuff. 556 00:32:00,810 --> 00:32:02,280 They're believing it now. 557 00:32:02,280 --> 00:32:05,580 DNA is the stuff. 558 00:32:05,580 --> 00:32:07,460 But how does it work? 559 00:32:07,460 --> 00:32:10,840 How can this dumb molecule possibly be the 560 00:32:10,840 --> 00:32:13,050 transforming principle? 561 00:32:13,050 --> 00:32:16,130 Well, smart, young people want to know. 562 00:32:18,750 --> 00:32:29,200 So an erstwhile ornithologist, that is a college kid from the 563 00:32:29,200 --> 00:32:33,220 University of Indiana who particularly liked bird 564 00:32:33,220 --> 00:32:37,490 watching got very enamored by this problem, actually based 565 00:32:37,490 --> 00:32:40,390 on some fabulous faculty at the University of Indiana. 566 00:32:40,390 --> 00:32:44,150 He got really intrigued by how could DNA possibly do this. 567 00:32:44,150 --> 00:32:47,600 But he recognized he didn't know any chemistry. 568 00:32:47,600 --> 00:32:53,010 He decided to go to Cambridge, England to the Medical 569 00:32:53,010 --> 00:32:56,440 Research Council lab, the MRC lab in Cambridge, England 570 00:32:56,440 --> 00:32:59,190 where he teamed up with someone who did a lot of 571 00:32:59,190 --> 00:33:02,350 talking and very few experiments. 572 00:33:02,350 --> 00:33:05,820 A physicist who had worked for the British admiralty during 573 00:33:05,820 --> 00:33:09,330 World War II on classified things and had somehow gotten 574 00:33:09,330 --> 00:33:12,440 interested in biology. 575 00:33:12,440 --> 00:33:15,110 And because he had this kid, this recently graduated 576 00:33:15,110 --> 00:33:19,310 college kid, and you had this 35-year-old physicist who 577 00:33:19,310 --> 00:33:22,840 nobody was quite sure what to make of, they kind of hung out 578 00:33:22,840 --> 00:33:24,760 with each other in the same office. 579 00:33:24,760 --> 00:33:27,960 And they didn't really do many experiments, but boy did they 580 00:33:27,960 --> 00:33:31,400 do a lot of talking, and thinking, and looking at all 581 00:33:31,400 --> 00:33:33,870 the data that were out there. 582 00:33:33,870 --> 00:33:37,510 And that's pretty much what James Watson and Francis Crick 583 00:33:37,510 --> 00:33:38,610 were doing. 584 00:33:38,610 --> 00:33:41,430 They knew this problem was important. 585 00:33:41,430 --> 00:33:44,330 And Jim and Francis would talk every day about this stuff, 586 00:33:44,330 --> 00:33:47,050 and they will talk to people down the hall who really knew 587 00:33:47,050 --> 00:33:49,290 about the chemistry of nucleic acids. 588 00:33:49,290 --> 00:33:55,230 And they went down to London to Maurice Wilkins' lab where 589 00:33:55,230 --> 00:33:57,660 crystals were being made of DNA. 590 00:33:57,660 --> 00:34:01,810 And Rosalind Franklin, who was a fantastic scientist and had 591 00:34:01,810 --> 00:34:05,930 managed to make crystals of DNA, showed Crick and Watson 592 00:34:05,930 --> 00:34:09,460 her crystals of DNA. 593 00:34:09,460 --> 00:34:11,670 Francis Crick being a physicist was very good at 594 00:34:11,670 --> 00:34:15,500 understanding crystallography and how crystal structures and 595 00:34:15,500 --> 00:34:17,949 x-ray diffraction patterns related to each other. 596 00:34:17,949 --> 00:34:22,550 And Francis knew immediately this thing had to be a helix. 597 00:34:22,550 --> 00:34:23,839 He could tell it was a helix. 598 00:34:26,380 --> 00:34:29,969 And they went back, and based on Rosalind Franklin's x-ray 599 00:34:29,969 --> 00:34:35,300 diffraction patterns, went and made a model, a model for the 600 00:34:35,300 --> 00:34:37,360 structure of DNA. 601 00:34:37,360 --> 00:34:39,330 You all know the model. 602 00:34:39,330 --> 00:34:41,389 You've seen the double helix forever. 603 00:34:41,389 --> 00:34:45,290 It's a cultural icon, but that's how this came about. 604 00:34:45,290 --> 00:34:57,530 And The Double Helix, the Structure of DNA, April of 605 00:34:57,530 --> 00:35:01,330 1953 is published. 606 00:35:01,330 --> 00:35:13,090 The double helix has two strands running an 607 00:35:13,090 --> 00:35:19,730 anti-parallel directions, 5 prime to 3 prime, 5 prime to 3 608 00:35:19,730 --> 00:35:25,950 prime anti-parallel directions, and it has a 609 00:35:25,950 --> 00:35:32,240 perfect base pairing between purines and pyrimidines. 610 00:35:32,240 --> 00:35:34,350 If you have a T, and I'm just going to 611 00:35:34,350 --> 00:35:37,050 draw this very quickly. 612 00:35:37,050 --> 00:35:39,620 You can look in your book for getting it just right. 613 00:35:39,620 --> 00:35:44,990 You have two hydrogen bonds. 614 00:35:57,340 --> 00:36:11,590 That's T and A, and if you have a C, you have three 615 00:36:11,590 --> 00:36:25,075 hydrogen bonds that perfectly hold it to the G, et cetera. 616 00:36:27,630 --> 00:36:32,520 So notice C and G fit perfectly together to make 617 00:36:32,520 --> 00:36:34,140 three hydrogen bonds. 618 00:36:34,140 --> 00:36:38,620 A and T fit perfectly together to make two hydrogen bonds, 619 00:36:38,620 --> 00:36:40,500 and that was the key was to recognize that when you stick 620 00:36:40,500 --> 00:36:43,140 them together in that way, you get exactly the same width. 621 00:36:43,140 --> 00:36:44,510 They fit perfectly. 622 00:36:44,510 --> 00:36:49,900 You couldn't match an A with a G, an A with a C, you could 623 00:36:49,900 --> 00:36:54,700 only match the A with a T. That was it. 624 00:36:54,700 --> 00:36:55,190 Brilliant. 625 00:36:55,190 --> 00:36:55,640 Beautiful. 626 00:36:55,640 --> 00:36:59,200 Now, you guys should read Crick's book The Double Helix 627 00:36:59,200 --> 00:37:01,610 in which he tells the stories because it's just a 628 00:37:01,610 --> 00:37:03,770 fascinating, fascinating business. 629 00:37:03,770 --> 00:37:06,900 He'll tell, or others will tell actually, the story. 630 00:37:06,900 --> 00:37:08,770 So you know what this means by the way? 631 00:37:08,770 --> 00:37:11,350 This means that the amount of A should equal the amount of 632 00:37:11,350 --> 00:37:14,470 T. And the amount of G should equal the amount of C. 633 00:37:14,470 --> 00:37:19,575 There should be a ratio, a one-to-one ratio that the A to 634 00:37:19,575 --> 00:37:23,230 T ratio should be one to one. 635 00:37:23,230 --> 00:37:26,520 And the G to C ratio should be one to one. 636 00:37:26,520 --> 00:37:30,340 Although any organism might have more As than Ts and Gs 637 00:37:30,340 --> 00:37:33,420 than Cs, the ratio of these guys should be one and these 638 00:37:33,420 --> 00:37:35,020 guys should be one. 639 00:37:35,020 --> 00:37:37,700 This actually was discovered by a chemist at Columbia 640 00:37:37,700 --> 00:37:38,950 called Chargaff. 641 00:37:41,640 --> 00:37:44,380 These were called Chargaff's rules. 642 00:37:44,380 --> 00:37:47,250 Chargaff was a very distinguished chemist who came 643 00:37:47,250 --> 00:37:50,036 up with Chargaff's rules with the As equals the Ts, and the 644 00:37:50,036 --> 00:37:51,720 Gs equals the Cs, and he didn't know 645 00:37:51,720 --> 00:37:53,760 what to make of it. 646 00:37:53,760 --> 00:37:57,920 By the way, Chargaff actually visited Cambridge, England 647 00:37:57,920 --> 00:38:00,700 while Crick and Watson were there before their discovery, 648 00:38:00,700 --> 00:38:02,630 and he had lunch with them. 649 00:38:02,630 --> 00:38:05,920 And he related that Crick and Watson seemed like Bozos to 650 00:38:05,920 --> 00:38:09,660 him, because they couldn't even keep straight the exact 651 00:38:09,660 --> 00:38:11,610 structure of the four bases. 652 00:38:11,610 --> 00:38:12,860 They always had to keep looking it up. 653 00:38:12,860 --> 00:38:15,180 They hadn't memorized the structures the four bases, and 654 00:38:15,180 --> 00:38:18,580 Chargaff was such a brilliant chemist, he, of course, knew 655 00:38:18,580 --> 00:38:20,270 this instantly, et cetera, et cetera. 656 00:38:20,270 --> 00:38:23,680 And he said, these guys are never going to get anywhere 657 00:38:23,680 --> 00:38:25,705 because they really don't even understand the structure of 658 00:38:25,705 --> 00:38:29,090 the bases, haven't memorized it. 659 00:38:29,090 --> 00:38:31,020 When Crick and Watson's discovery turned out to be the 660 00:38:31,020 --> 00:38:32,940 single most important biological discovery of the 661 00:38:32,940 --> 00:38:37,770 20th century, Erwin Chargaff who lived a very long life was 662 00:38:37,770 --> 00:38:41,340 sort of bitter because he's kind of worked it out in a way 663 00:38:41,340 --> 00:38:44,680 with the ratio and never figured out what it meant. 664 00:38:44,680 --> 00:38:46,835 And he said one of the bitterest, cuttingest comments 665 00:38:46,835 --> 00:38:51,220 I've have ever heard from a scientist which is referring 666 00:38:51,220 --> 00:38:53,850 to Crick and Watson as still not being impressed even after 667 00:38:53,850 --> 00:38:55,120 they won their Nobel Prize for this. 668 00:38:55,120 --> 00:39:00,940 He said that two such pygmies should cast such giant shadows 669 00:39:00,940 --> 00:39:03,560 only shows how late in the day it is. 670 00:39:06,490 --> 00:39:09,810 Anyway, he was not happy to have missed this point. 671 00:39:09,810 --> 00:39:11,840 Crick and Watson were very happy to have 672 00:39:11,840 --> 00:39:13,180 figured this out. 673 00:39:13,180 --> 00:39:17,330 They, when they figured this out in February of 1953, what 674 00:39:17,330 --> 00:39:18,710 do you do in England when you make a big discovery? 675 00:39:18,710 --> 00:39:20,596 AUDIENCE: You have tea. 676 00:39:20,596 --> 00:39:23,284 PROFESSOR: No, you don't have tea. 677 00:39:23,284 --> 00:39:25,140 You go to the pub. 678 00:39:25,140 --> 00:39:27,560 They went to the pub. 679 00:39:27,560 --> 00:39:30,480 They ran down to the Eagle Pub, they brought everybody 680 00:39:30,480 --> 00:39:32,780 drinks, and they told everybody at the Eagle Pub, 681 00:39:32,780 --> 00:39:35,750 we've discovered the secret of life. 682 00:39:35,750 --> 00:39:37,590 The people at the Eagle Pub had no idea what they were 683 00:39:37,590 --> 00:39:40,980 talking about, but were happy to have a round of drinks, and 684 00:39:40,980 --> 00:39:42,190 there you go. 685 00:39:42,190 --> 00:39:45,660 They immediately raced to write this up in Nature. 686 00:39:45,660 --> 00:39:49,290 It appears in Nature in April of 1953, and 687 00:39:49,290 --> 00:39:50,670 it's a one-page paper. 688 00:39:50,670 --> 00:39:52,360 And get it on the web and read it. 689 00:39:52,360 --> 00:39:55,490 It's the single best one page that has been written in 690 00:39:55,490 --> 00:39:58,130 biology in the 20th century. 691 00:39:58,130 --> 00:39:59,515 And of course, what did they say? 692 00:40:02,400 --> 00:40:05,590 The title is kind of unassuming, "A Structure for 693 00:40:05,590 --> 00:40:09,360 the Salt of Deoxyribonucleic Acid," nothing too exciting. 694 00:40:09,360 --> 00:40:11,460 But Crick and Watson realized something. 695 00:40:11,460 --> 00:40:13,290 What do they realize? 696 00:40:13,290 --> 00:40:18,440 Crick and Watson realized why is this a big deal? 697 00:40:18,440 --> 00:40:21,320 Why is this double helix so important? 698 00:40:21,320 --> 00:40:38,850 Well, the implication of the double helix is that if I have 699 00:40:38,850 --> 00:40:47,680 a double helix and those strands were to separate, each 700 00:40:47,680 --> 00:40:56,930 would be a template for a double helix, heredity. 701 00:40:56,930 --> 00:41:00,660 How do you pass information to two daughter cells? 702 00:41:00,660 --> 00:41:01,700 You got a double helix. 703 00:41:01,700 --> 00:41:03,200 It's redundant. 704 00:41:03,200 --> 00:41:04,830 If you know the A is on one strand, you know 705 00:41:04,830 --> 00:41:07,230 the Ts on the other. 706 00:41:07,230 --> 00:41:11,710 Unzip it, copy, voila. 707 00:41:11,710 --> 00:41:14,020 I now have two copies of heredity. 708 00:41:14,020 --> 00:41:15,270 What's a mutation? 709 00:41:18,240 --> 00:41:21,310 Occasionally get it wrong. 710 00:41:21,310 --> 00:41:21,910 Bingo. 711 00:41:21,910 --> 00:41:22,770 They saw it. 712 00:41:22,770 --> 00:41:24,020 They knew. 713 00:41:26,612 --> 00:41:29,390 Now, they didn't have time to prove this. 714 00:41:29,390 --> 00:41:30,960 I mean, who has time to prove this. 715 00:41:30,960 --> 00:41:32,940 This is such an exciting discovery secret of life. 716 00:41:32,940 --> 00:41:36,580 Drinks for everybody, they write it up, but in the last 717 00:41:36,580 --> 00:41:39,520 paragraph, they certainly don't want anybody to think 718 00:41:39,520 --> 00:41:41,110 that they missed the point. 719 00:41:41,110 --> 00:41:45,160 And they write the coyest sentence in molecular biology. 720 00:41:45,160 --> 00:41:50,250 They write, "It has not escaped our notice that this 721 00:41:50,250 --> 00:41:52,480 structure offers an explanation for 722 00:41:52,480 --> 00:41:54,580 heredity and mutation. 723 00:41:54,580 --> 00:41:59,870 We'll address this in another paper." Cute. 724 00:41:59,870 --> 00:42:01,130 Very cute. 725 00:42:01,130 --> 00:42:04,290 They put down their marker, they knew what it meant, but 726 00:42:04,290 --> 00:42:06,720 they got the thing off to Nature very quickly. 727 00:42:06,720 --> 00:42:07,680 It was a hot topic. 728 00:42:07,680 --> 00:42:08,950 They were competing with other people. 729 00:42:08,950 --> 00:42:11,030 You'll read about the competition with Linus Pauling 730 00:42:11,030 --> 00:42:12,550 and other things like that. 731 00:42:12,550 --> 00:42:15,030 It had not escaped their notice that this pretty much 732 00:42:15,030 --> 00:42:17,410 explains heredity. 733 00:42:17,410 --> 00:42:18,860 Now, of course, are you convinced 734 00:42:18,860 --> 00:42:20,330 that it explains heredity? 735 00:42:20,330 --> 00:42:24,480 It's a nice model, but don't we require proof? 736 00:42:24,480 --> 00:42:25,520 We do require proof. 737 00:42:25,520 --> 00:42:27,800 It's an ex post facto model, we need proof, It's a pretty 738 00:42:27,800 --> 00:42:31,230 good ex post facto model, but we need proof. 739 00:42:31,230 --> 00:42:39,650 So the last step, which I'll touch on very briefly, was 740 00:42:39,650 --> 00:42:46,530 proof of what's called semi-conservative replication. 741 00:42:52,300 --> 00:42:56,720 Meaning that each strand is used for the other. 742 00:42:56,720 --> 00:42:58,210 And I might run two minutes over. 743 00:42:58,210 --> 00:42:58,640 We'll see. 744 00:42:58,640 --> 00:43:00,310 I'll try to keep it within time. 745 00:43:03,900 --> 00:43:08,290 Out at Caltech, two graduate students, Frank Stahl and Matt 746 00:43:08,290 --> 00:43:11,020 Meselson hear of this. 747 00:43:11,020 --> 00:43:12,890 Matt Meselson by the way is still 748 00:43:12,890 --> 00:43:14,370 working in Harvard Square. 749 00:43:14,370 --> 00:43:15,120 He's at Harvard. 750 00:43:15,120 --> 00:43:16,520 He's a wonderful guy. 751 00:43:16,520 --> 00:43:17,130 Matt is there. 752 00:43:17,130 --> 00:43:19,410 You could ask Matt about this, young graduate student at 753 00:43:19,410 --> 00:43:21,590 Caltech in the early '50s. 754 00:43:21,590 --> 00:43:24,660 Obviously, this model looks like it must be right. 755 00:43:24,660 --> 00:43:27,170 How do you prove it? 756 00:43:27,170 --> 00:43:33,830 Well, Matt and Frank, Meselson and Stahl came up with a cool 757 00:43:33,830 --> 00:43:35,690 experiment to prove it. 758 00:43:35,690 --> 00:43:46,940 Meselson and Stahl, they take bacteria, here's my DNA in 759 00:43:46,940 --> 00:44:01,320 there, they want to show that each strand, each, when we 760 00:44:01,320 --> 00:44:06,170 make a new generation of bacteria, the new DNA has one 761 00:44:06,170 --> 00:44:09,270 old strand and one new strand. 762 00:44:09,270 --> 00:44:11,400 That each old strand is being used as a 763 00:44:11,400 --> 00:44:13,200 template for a new strand. 764 00:44:13,200 --> 00:44:16,240 How are we going to tell? 765 00:44:16,240 --> 00:44:17,680 We gotta label it somehow. 766 00:44:17,680 --> 00:44:20,875 We got a label the old strand different than the new strand. 767 00:44:23,710 --> 00:44:25,880 How do we possibly label and they're the same chemical 768 00:44:25,880 --> 00:44:26,930 composition? 769 00:44:26,930 --> 00:44:28,115 What are we going to do? 770 00:44:28,115 --> 00:44:28,985 AUDIENCE: Radioactivity. 771 00:44:28,985 --> 00:44:33,600 PROFESSOR: Radioactivity or some kind of isotope. 772 00:44:33,600 --> 00:44:35,950 Well, they used an isotope. 773 00:44:35,950 --> 00:44:40,660 What they did, super cool, they grew this up, but not in 774 00:44:40,660 --> 00:44:46,540 normal nitrogen, but in N-15, not, by the way, radioactive, 775 00:44:46,540 --> 00:44:47,780 but different weight. 776 00:44:47,780 --> 00:44:50,560 They grew it up in N-15. 777 00:44:50,560 --> 00:44:54,000 They added bacterias that had grown up, and all of their DNA 778 00:44:54,000 --> 00:44:58,240 had lots of N-15 in it on both strands. 779 00:44:58,240 --> 00:45:03,760 They then pour in a lot of medium that has N-14. 780 00:45:03,760 --> 00:45:06,650 Tons, they swamp it with N-14 so what's this 781 00:45:06,650 --> 00:45:08,610 strand going to be? 782 00:45:08,610 --> 00:45:12,520 N-14. 783 00:45:12,520 --> 00:45:17,660 So what can you tell me about the difference between this 784 00:45:17,660 --> 00:45:19,692 DNA and that DNA? 785 00:45:19,692 --> 00:45:21,420 AUDIENCE: One's going to be lighter. 786 00:45:21,420 --> 00:45:23,430 PROFESSOR: One is going to be lighter. 787 00:45:23,430 --> 00:45:26,800 How do you measure how much lighter it is? 788 00:45:26,800 --> 00:45:31,920 They came up, they invented the technique for this purpose 789 00:45:31,920 --> 00:45:34,920 of centrifuging in a salt gradient. 790 00:45:34,920 --> 00:45:39,860 They put in the right amounts of salt, cesium salt, and they 791 00:45:39,860 --> 00:45:43,120 centrifuge it so hard that there's a gradient of 792 00:45:43,120 --> 00:45:46,730 densities, denser here, lighter here. 793 00:45:46,730 --> 00:45:54,570 And they find that this DNA and this DNA centrifuge to 794 00:45:54,570 --> 00:45:56,900 different places. 795 00:45:56,900 --> 00:45:59,300 There's a difference in their density. 796 00:45:59,300 --> 00:46:05,300 The new DNA is half old, N-15, half new, N-14. 797 00:46:05,300 --> 00:46:06,810 It has the intermediate density. 798 00:46:06,810 --> 00:46:11,060 If I grow another generation, I'm going to see some N-15 799 00:46:11,060 --> 00:46:15,240 14s, and I'm also going to see some N-14, 14s. 800 00:46:15,240 --> 00:46:18,050 And that's what they found. 801 00:46:18,050 --> 00:46:22,110 They invented this technique it's isopycnic centrifugation 802 00:46:22,110 --> 00:46:27,790 and Matt Meselson and Frank Stahl provided an experimental 803 00:46:27,790 --> 00:46:32,940 prediction of the beautiful Crick-Watson model that was 804 00:46:32,940 --> 00:46:36,170 fair to say the secret of life. 805 00:46:36,170 --> 00:46:39,200 Anyway, these are the foundations. 806 00:46:39,200 --> 00:46:43,560 Notice now, we've taken genetics and used it to do 807 00:46:43,560 --> 00:46:44,840 biochemistry. 808 00:46:44,840 --> 00:46:47,500 We've taken biochemistry and used it 809 00:46:47,500 --> 00:46:50,230 to understand genetics. 810 00:46:50,230 --> 00:46:54,590 And so finally, we are making the bridge 811 00:46:54,590 --> 00:46:56,580 of molecular biology. 812 00:46:56,580 --> 00:46:57,830 Next time.