The mode of DNA replication was determined in 1958 by Meselson and Stahl. They designed an experiment to distinguish between semiconservative, conservative and dispersive replications. In their experiment, they grew two cultures of E.coli for many generations in separate media. The ‘heavy’ culture was grown in a medium in which the nitrogen source (NH4CI) contained the heavy isotope 15N and the ‘ light’ culture was grown in a medium in which the nitrogen source contained light isotope 14H for many generations. At the end of growth, they observed that the bacterial DNA in the heavy culture contained only 15N and in the light culture only 14N. The heavy DNA could be distinguished from light DNA ( 15N from 14N) with a technique called Cesium Chloride (CsCl) density gradient centrifugation. In this process, heavy and light DNA extracted from cells in thtytwo cultures settled into two distinct and separate bands (hybrid DNA).
The heavy culture ( 15N) was then transferred into a medium that had only NH4CI, and took samples at various definite time intervals (20 minutes duration). After the first replication, they extracted DNA and subjected it to density gradient centrifugation. The DNA settled into a band that was intermediate in position between the previously determined heavy and light bands. After the second replication (40 minutes duration), they again extracted DNA samples,and this time found the DNA settling into two bands, one at the light band position and one at intermediate position. These results confirm Watson and Crick’s semi – conservative replication hypothesis.
