Hi all,
I am doing a genescan analysis for a deleted exon in the patient sample. The amplified region is 215 bp. In the patient sample because of homozygous deletion, no peak is observed while in control I am getting three peaks of sizes 215, 216 and 217bp. Is there any possible explanation for this? I have attached the results for reference.
PS: I use albumin as control and getting a single peak as expected. The reverse primer after UCSC BLAT gives more than one hit while the forward gives only one hit.
Since the peaks are +/- 1base it may be that your amplified template has a polyA or other long string of a single base and that you have enzyme slippage in the amplification generating different lengths of the polynucleotide motif. You can get a +1 band from taq polymerase adding an A base at the end of an amplimer generally but as you control peak does not show this effect it is unlikely that this mechanism is contributing a peak in your deletion amplimer.
It might just be possible if the poly A tract exists in the 215 and 216 sizes that the 217 is a heteroduplex of 215+216 moving slowly through the capillary but it does not seem very likely
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