Reminder on RFLP and SSR analysis (Background)

The boys in Implementation and Planning keep whining about not understanding the jargon we keep using, so I had an intern write up a short summary of the more mundane technology we're talking about. If you have questions or complaints, just e-mail me and I'll be sure the little twerp gets an F. If it's really bad, then I'll just have Bill register him as a sex offender after we send him back to college.

Humans are a diploid mammalian eukaryote, with 22 pairs of autosomes and 1 pair of sex chromosomes, with gender determined by Y-mediated “XY” determination. At present estimate, humans have approximately 20,000-25,000 genes, with only about 1.5% of the total length serving as protein-coding exons, with the rest being occupied by transposons, pseudo-genes, regulatory elements, tandem and interspersed repeats, as well as other, undiscovered genetic elements.

All genetic analysis relies on a technique called Polymerase Chain Reaction (PCR), which is used to amplify (exponentially copy) small samples of DNA. The section of DNA amplified can be targeted, since the DNA needs an RNA primer to begin copying. If the sequence of nucleotides flanking the gene or segment of interest is known, then RNA templates can be constructed that anneal to the flanking sequence.

When these primers, the DNA of interest, and taq DNA polymerase (the protein responsible for copying DNA. Taq DNA polymerase is a polymerase extracted from a thermophillic Archaebacteria found in the geysers of Yellowstone, and thus are not damaged by high temperatures) are mixed together, melting the DNA allows the RNA templates to bind to the flanking sequences, which allows the taq DNA polymerase to begin copying both strands of DNA. This produces two strands of DNA, both of which have one molecule of original DNA, and one molecule of newly created DNA. This process is repeated several dozen times, at which point, you have (2n - 2) molecules of new DNA, and 2 molecules of original DNA. This process can be altered in many ways allowing for sequencing of the DNA, copying of multiple sites, as well as finding out the rate at which the DNA reanneals or melts, to find the repetitiveness of the DNA, or the ratio of A+T:G+C composition, respectively.