10 How many copies of a gene
So how do we know how many copies of a gene we had at the start?
Consider how the number of copies of DNA that is present in the initial reaction tube will influence how many copies of amplified DNA are present at each cycle of the reaction.
If you start at cycle 1 with 2 copies, then by 20 cycles there are just over 4 million copies.
How many copies would there at 20 cycles if you had started with 4 copies?
Answer
There would be twice as many, over 8 million, as you would have twice as many to start from and each stage duplicates each DNA molecule.
We can use this idea in PCR to work out the number of starting copies present. The PCR machine automatically records when any particular test sample crosses a threshold value of fluorescence (amount of green light) that is detected in a tube. This can then be compared with positive control samples that have known copy numbers of the CYP2D6 gene or of particular variants. In this way we can work out how many copies of the CYP2D6 gene are present in that individual’s genome.
In order to make this comparison we need to ensure that every sample that is analysed contains the same amount of genomic DNA (and hence the same number of copies of chromosome 22 carrying the CYP2D6 gene).
9 The PCR machine