DNA replication

DNA replication The mode of DNA replication was determined in by Meselson and Stahl. They designed an experiment to distinguish between semi conservative, 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 (NH Cl) contained the heavy isotope N and the ‘light’ culture was grown in a medium in which the nitrogen source contained light N-DNA Generation I Generation II Gravitational force Heavy Hybrid Light Hybrid N-DNA N N N N N N N N N-DNA min min N-DNA N-DNA N-DNA N-DNA Fig. . Meselson and Stahl experiment to support semiconservative mode of DNAreplication XII Std Biology-Zoology Chapter- XII Std Biology-Zoology Chapter- Molecular Genetics isotope N for many generations. At the end of growth, they observed that the bacterial DNA in the heavy culture contained only N and in the light culture only N. The heavy DNA could be distinguished from light DNA ( N from N) with a technique called Cesium Chloride (CsCl) density gradient centrifugation . In this process, heavy and light DNA extracted from cells in the two cultures settled into two distinct and separate bands (hybrid DNA) ( Fig. . ). The heavy culture ( N) was then transferred into a medium that had only NH Cl, and took samples at various definite time intervals ( 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 ( 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.