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SEMICONSERVATIVE MODE OF DNA REPLICATION In 1958 Matthew Meselson and Franklin Stahl performed an experiment that confirmed the

semiconservative nature of DNA replication. They grew cultures of E. coli for many generations on media in which the only source of nitrogen was the heavy isotope 15N. After several generations of growth on heavy-isotope medium, essentially all the nitrogen atoms in the DNA of these bacterial cells were labeled with (that is, contained) 15N. When DNA is extracted from these cells and centrifuged on CsCl density gradient centrifugation, the DNA separates out at the point at which its density equals that of the salt solution. The cells in some of these cultures were then transferred to new medium in which all the nitrogen was 14N. Any DNA synthesized after the transfer would contain the lighter isotope. DNA was extracted periodically and was compared to pure 14N DNA and 15N DNA. After one round of replication, the resultant DNA was found to have close to the intermediate density. Since conservative replication would result in equal amounts of DNA of the higher and lower densities (but no DNA of an intermediate density), conservative replication was excluded. However, this result was consistent with both semi-conservative and dispersive replication. Semi-conservative replication would result in doublestranded DNA with one strand of 15N DNA, and one of 14N DNA, while dispersive replication would result in double-stranded DNA with both strands having mixtures of 15N and 14N DNA. Either of these possibilities would have resulted in appearance of DNA of an intermediate density. The authors continued to sample cells as replication continued. DNA from cells after two replications had been completed was found to consist of equal amounts of DNA with two different densities, one corresponding to the intermediate density of DNA of cells grown for only one division in 14N medium, the other corresponding to DNA from cells grown exclusively in 14N medium. This was inconsistent with dispersive replication, which would have resulted in a single density, lower than the intermediate density of the one-generation cells, but still higher than cells grown only in 14N DNA medium, as the original 15 N DNA would have been split evenly among all DNA strands. By confirming the predictions of semi-conservative replication, the Meselson-Stahl experiment disproved the conservative and dispersive alternatives. We now know that the semi-conservative replication of DNA is nearly universal.

Concept Check
Q1. The best adjective to describe DNA replication is (a).nondisruptive. (b) semi-conservative. (c) conservative. (d) dispersive

Q2.In the Meselson-Stahl experiment, which of the three modes of replication could be ruled out (a) after one round of replication? (b) after two rounds of replication? Explain why. Q3. Describe the role of N
15

in the Meselson-Stahl experiment.


14

Q4. (i) In a semi-conservative DNA replication model, what would be the distribution of N molecules after (a) first generation of growth in N
14

and N

15

in the DNA

media and (b) second generation of growth?

(ii) Work out the above question by assuming conservative model of DNA replication.