Insofar as I have any “philosophy” about how I do my work, it’s this: keep experiments simple. I’ve always tried to do experiments sufficiently uncomplicated and easy to understand that the results—one way or the other—would be clear-cut enough to not require (or barely require) statistical analysis. I’ve taught my students this notion, too, and I think I’ve succeeded in that endeavor.
And the experiments I most admire are equally simple. The most beautiful, as I’ve mentioned before, is Meselson and Stahl’s demonstration, in 1958, that the replication of DNA was “semiconservative”: that is, when a two-stranded DNA molecule replicates, it unzips and each strand forms a template for building a new strand from nucelotide and sugar constituents. (There are many other ways DNA could have replicated.)
The results of this experiment were crystal clear: they involved first marking all the DNA strands in bacteria with a heavy isotope of nitrogen (you can do this simply by giving the bacteria Purina’s heavy-nitrogen E. Coli Chow). They then put those bacteria into medium that had “light” nitrogen, extracting the bacterial DNA at various intervals—as their DNA replicated—and centrifuging it in a cesium chloride density gradient. This enables you to see which strands have the heavy nitrogen and which don’t, for the different-weight strands move to different positions in the chemical gradient; and various theories of how DNA replicates predict different patterns for how the strands will migrate. I’d recommend simply downloading the Meselson and Stahl paper at the link below to see how clean the results were. You don’t have to be a scientist to understand how the experiment worked, and what it means. (This, and the experiment following, are all described beautifully in Horace Freeland Judson’s The Eighth Day of Creation.)
Below is figure 4 from Meselson and Stahl’s paper, which shows absolutely, and without any need for mathematical analysis, that DNA replicates semiconservatively. See how a “heavy” (high-weight) band starts giving rise to a lighter band within one generation (a DNA molecular that’s half the original “light” one and half the new “heavy”one). Lighter-weight bands are to the left. And then, after another generation of DNA replication (the bacterium E. coli replicates every 20 minutes), you see even lighter bands, now consisting of the newly formed light strands which have themselves become templates for yet another light strand—giving rise to “double light” DNA. Read from the top down: from the beginning of the experiment to the end, note how heavier bands produce semi-heavy bands (half old, half new DNA) and then fully light bands (all new DNA).
Lighter-weight bands are to the left:
This is what scientists call “a clean result”
The second most beautiful experiment, which took place fifty years and one week ago, was done by Marshall Nirenberg and J. Heinrich Matthaei, and is the subject of a really nice article by our pinch-“blogger” Matthew Cobb in yesterday’s Telegraph “Genes and DNA: meet the first man to read the book of life.” The piece is really about Matthaei, who did the crucial experiment, in 1961, that began the decoding of DNA. By “decoding”, I mean understanding how the sequence of four nucleotide “letters” in DNA (adenine, guanine, cytosine, and thymine [which is “uracil” in the RNA product produced by DNA]) codes for amino acids, the constituents of proteins. After all, what DNA “does”, by and large, is code for proteins.
It had been theorized by George Gamow that because there are 4 DNA bases and 20 amino acids, the code was probably a triplet code, since with 4 bases a doublet code could only yield 16 (4 X 4) amino acids. Unravelling this code was one of the major accomplishments of modern biology, and was begun by Nirenberg and Matthaei in Nirenberg’s lab at the National Institutes of Health. As Matthew describes, the crucial experiment was actually done by Matthaei while Nirem=nberg was away. Setting it up was complicated, for it required constructing a system of “cell-free” protein synthesis, made by using the cell contents of bacteria. Once in place, the researchers could use artificially constructed RNAs (the product of DNA that itself codes for proteins) to see what proteins could be produced by RNA in the artificial system.
As described on pp. 473-480 in The Eighth Day of Creation (buy that book!), the crucial experiment, called “27Q,” was begun at 3 a.m. (!) on May 27, 1961. It was over six hours later. Matthaei determined that an artificial strand of RNA composed only of the nucleotide base uracil (“poly-U”) produced proteins containing only phenylalanine. Thus “UUU” (or “UUUU” or higher polymers; they didn’t yet know the code was triplet) coded for that amino acid. They cracked codes for other amino acids as well. As Matthew describes in his piece:
The discovery was finally revealed two weeks later in Moscow, at the Fifth International Congress of Biochemistry. Nirenberg was given 15 minutes to present his findings – but only a handful of people turned up to hear a nobody claim he had solved a problem that was still defeating the world’s largest laboratories. When the news reached Francis Crick that afternoon, he immediately changed the conference programme so that the young American could give his talk again. The next day, in front of a packed lecture theatre, Nirenberg described his careful experiments and created a sensation. When he stepped on to the stage, Nirenberg also stepped into history.
By the end of the year, Crick had shown that the DNA code was a triplet code, and that code had been cracked for every amino acid.
Nirenberg stepped into history, but Matthaei only got his toe in. For in 1968 the Nobel Prize for Medicine and Physiology was given to Robert Holley, H. Gobind Khorana, and Marshall W. Nirenberg for uncovering the relationship between DNA sequences and proteins. Matthaei was left out in the cold. (Nobel Prizes in one area cannot be given to more than three people in a given year). This is quite unjust, since Matthaei had done the crucial experiment and was also pivotal in setting up the cell-free synthesis system. As usual, the boss gets the prizes and the grunts get squat. I consider Meselson and Stahl’s lack of Nobels equally unjust.
Nirenberg died last year, but what happened to Matthaei? Surprisingly, even at age 82 he still goes to the lab, bicycling to the Max-Planck Institute every day. You can see a video of him and an interview (in German) here. What a trouper!
Nirenberg, M. W., and J. H. Matthaei. 1961. The dependence of cell-free protein synthesis in E. coli upon naturally occurring or synthetic polyribonucleotides. Proc Natl Acad Sci U S A 47:1588-602.