Today we extracted DNA from our cheek cells, and then set up a PCR (Polymerase Chain Reaction) targeting the PV92 locus.
PV92 is found in chromosome 16 in humans. We all have two copies of each chromosome, so we have two chromosomes 16, and therefore we have two copies of the PV92 locus.
What does PV92 do? Nothing! It does not code for any thing, just as the 95% of the human genome. Only about 5% of our genome actually encodes some sort of product. This locus was named in the old days of karyiotyping, when we didn't really know what different areas stained in a chromosome did. After we sequenced the human genome, we figured out that the names given to many loci didn't actually correspond to a functional gene. For practical purposes we'll consider PV92 a gene, with a couple of "alleles" (see below).
The nice thing of having a locus that doesn't do any thing at all is that it is not influenced by
natural selection. So when we think about mating in primate populations, PV92 reflects the mating patterns, since the allele frequency is not going to be incluenced by selection. So? Well... in population genetics a lot of hypothesis testing is based on the premise of
random mating in a given population. PV92 has been used for a lot of human population genetics studies.
There is another nice thing about PV92. Some people have an
Alu sequence inserted in the very middle of the PV92 locus. This Alu insert (which could be an intron should PV92 be a real gene. An intron is a DNA fragment found in between coding regions of a gene, that doesn't code for any thing and that it's spliced during transcription) makes PV92 300 bases longer, which is a difference big enough to be visualized in an agarose gel (originally PV92 is about 600 base pairs long).
An Alu sequence is a retrotransposon (a DNA fragment that has been inserted in a location where it doesn't belong via
reverse transcription,
i.e. mRNA actually writes a sequence into the genome) found throughout primate genomes. There are thousands of copies of Alu sequences found in a single genome.
PCR, the
Polymerase Chain Reaction, a method to amplify (make many copies of) specific DNA fragments, is used to generate enough DNA that it can be studied with a variety of thechniques. Using PCR we will amplify our PV92 loci to see if we have the Alu sequence insert in any of our copies of PV92. Each one of us may have 0, 1, or 2 alu inserts in our PV92 copies and we will find out through PCR and gel electrophoresis.
Today we
extracted our own DNA. We scraped our cheeks to get some epithelial cells, we neutralized ions (with the InstaGel) that may be used as cofactors by
DNases to chop out DNA, and then we bursted our cells open (with the water baths) to extract the DNA from the nuclei.
We added our DNA, plus three positive controls, to a
master mix with all the constituents of a PCR: buffer, nucleotides,
Taq polymerase, magnesium, and primers (synthesized single stranded oligonucleotide sequences) and put the tubes into the thermocycler.
Tomorrow we'll run a gel electrophoresis to figure out who among us has the Alu insert in his/her PV92 locus, and if it is a single copy or two.
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