Semiconservative replication

Semiconservative replication describes the method by which DNA is replicated in all known cells.This method of replication was one of three proposed modelscite book |author=Anthony J. F. Griffiths et al. |title=An Introduction to genetic analysis |publisher=W.H. Freeman |location=San Francisco |year=1999 |chapter = 8. The Structure and Replication of DNA |chapterurl = http://www.ncbi.nlm.nih.gov/books/bv.fcgi?rid=iga.section.1505 |isbn=0-7167-3520-2] [Meselson, M. and Stahl, F.W. (1958). "The Replication of DNA in Escherichia coli". PNAS 44: 671-82. PMID 16590258.] of DNA replication:
* Conservative replication would leave the two original template DNA strands together in a double helix and would produce a copy composed of two new strands containing all of the new DNA base pairs.
* Dispersive replication would produce two copies of the DNA, both containing distinct regions of DNA composed of either both original strands or both new strands.
* Semiconservative replication would produce two copies that each contained one of the original strands and one entirely new strand.

The deciphering of the structure of DNA by Watson and Crick in 1953 suggested that each strand of the double helix would serve as a template for synthesis of a new strand. However, there was no way of guessing how the newly synthesized strands might combine with the template strands to form two double helical DNA molecules. The semiconservative model seemed most reasonable since it would allow each daughter strand to remain associated with its template strand. The semiconservative model was verified by the "Meselson-Stahl experiment" and other even more revealing experiments that allowed for autoradiographic visualization of the distribution of old and new strands within replicated chromosomes.

Testing the semi-conservative theory

The semi-conservative theory can be proven making use of the fact that DNA is made up of nitrogen bases. Nitrogen has an isotope N15 (N14 is the most common isotope) called heavy nitrogen. The experiment that proves the semi-conservative theory makes use of this isotope and runs as follows:

1.- Bacterial (E coli) DNA is placed in a media containing heavy nitrogen, which binds to the DNA, making it identifiable. 2.- This DNA is then placed in a media with the presence of N14 and left to replicate only once. The new bases will contain nitrogen 14 while the originals will contain N15 3.- The DNA is placed in test tubes containing caesium chloride (heavy compound) and centrifuged at 40,000 revolutions per hour. 4.- The caesium chloride molecules sink to the bottom of the test tubes creating a density gradient. The DNA molecules will position at their corresponding level of density (taking into account that N15 is more dense than N14)5.- These test tubes are observed under ultraviolet rays. DNA appears as a fine layer in the test tubes at different heights according to their density.

* According to the semi-conservative theory, after one replication of DNA, we should obtain 2 hybrid (part N14 part N15) molecules from each original strand of DNA. This would appear as a single line in the test tube. This result would be the same for the dispersive theory.
* On the other hand, according to the conservative theory, we should obtain one original DNA strand and a completely new one i.e. two fine lines in the test tube placed separately one from the other. Up to this point, either the semi-conservative or the dispersive theories could be truthful, as experimental evidence proved that only one line appeared after one replication. In order to conclude between those two, DNA had to be left to replicate again, still in a media containing N14.
* In the dispersive theory, after 2 divisions we should obtain a single line, but further up in the test tube, as the DNA molecules become less dense as N14 becomes more abundant in the molecule
* According to the semi-conservative theory, 2 hybrid molecules and 2 fully N14 molecules should be produced, so two fine lines at different heights in the test tubes should be observed. Experimental evidence proved that two lines were observed therefore offering compelling evidence for the semi-conservative theory.

ee also

*Molecular structure of Nucleic Acids

References