Course: SAT 170 Introductory Biotechnology

Unit 3 Reading and Multimedia Assignment

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Page 7 of 13
Attributions
Excerpt from “Biology: Boundless Biology Open Textbook”, CC BY SA 3.0

https://www.boundless.com/biology/dna/
Complementary Base Pairing

During DNA replication, each of the two strands of DNA serve as the template for the synthesis of a new, complementary strand.

The double-helix structure of DNA suggested to Watson and Crick the mechanism by which DNA, hence genes, could be copied faithfully. They proposed that when the time came for DNA to be replicated, the two strands of the molecule separated from each other, but remained intact, as each served as the template for the synthesis of a complementary strand. When the replication process is complete, two DNA molecules, which are identical to each other and identical to the original, have been produced. This mode of replication is described as the semiconservative model; one-half of each new molecule of DNA is old and one-half new (see figure above). In 1954, Watson and Crick published their hypothesis of the semiconservative DNA replication in a follow-up paper to their landmark publication of the DNA structure. While Watson and Crick had only suggested that this was the way the DNA would replicate, proof of the model came from the experiments of Matthew Meselson and Franklin Stahl.

In addition to Watson and Crick's semiconservative hypothesis, two other hypotheses were proposed for the method of DNA replication. The conservative hypothesis proposed that the entire DNA molecule acted as a template for the synthesis of an entirely new one. According to this model, histone proteins bound to the DNA, distorting it in such a way as to expose both strands' bases for hydrogen bonding. The dispersive hypothesis was exemplified by a model proposed by Max Delbrück. This model attempted to solve the problem of unwinding the two strands of the double helix by a mechanism that breaks the DNA backbone every 10 nucleotides or so, untwists the molecule, and then attaches the old strand to the end of the newly synthesized one. This would synthesize the DNA in short pieces alternating from one strand to the other.

Models for DNA Replication

Each of these three models made a different prediction about the distribution of the old DNA in molecules formed after replication. In the conservative hypothesis, after replication, one molecule is the entirely conserved old molecule, and the other is all newly synthesized DNA. The semiconservative hypothesis predicts that each molecule, after replication, will contain one old and one new strand. The dispersive model predicts that each strand of each new molecule will contain a mixture of old and new DNA. The three models are compared in the above figure.

Meselson-Stahl Experiment

Meselson and Stahl tested the viability of these hypotheses with an experiment (see figure above). They grew E. coli bacteria in a medium using ammonium ions (NH4+) as the source of nitrogen for DNA (as well as protein) synthesis. 14N is the common isotope of nitrogen, but Meselson and Stahl also used ammonium ions that were enriched for a rare heavy isotope of nitrogen, 15N. After growing E. colifor several generations in a medium containing 15NH4+, the researchers found that the DNA of the cells was heavier than normal because of the 15N atoms in it. The difference could be detected by extracting DNA from the E. coli cells and spinning it in an ultracentrifuge. Then Meselson and Stahl transferred more living cells that had been growing in 15NH4+ to a medium containing ordinary ammonium ions (14NH4+) and allowed them to divide just once. The DNA in this new generation of cells was exactly intermediate in density between that of the previous generation and the normal. This, in itself, is not surprising. It tells us no more than that half the nitrogen atoms in the new DNA are 14N and half are 15N. It tells us nothing about their arrangement in the molecules. However, when the bacteria were allowed to divide again in normal ammonium ions (14NH4+), two distinct densities of DNA were formed; half the DNA was normal and half was intermediate. These results showed that DNA molecules were not degraded and reformed from free nucleotides between cell divisions, but instead each original strand remained intact as it built a complementary strand from the nucleotides available to it. Meselson and Stahl's work provided definitive evidence for the semiconservative model of replication because each daughter DNA molecule is one-half old and one-half new.