I performed PCR amplification of genomic DNA isolated from human primary melanocytes.
I got no bands, positive control worked fine.
Was told by my PI that melanin produced by melanocyte impede polymerase activity during PCR (why he didn't tell me before i do not know...)
I found protocols online which require reisolation of DNA from cells.
A) I was wondering if anybody knows how I can inhibit melanin in my already isolated DNA samples.
I did find multiple papers suggesting 'blocking' with BSA by adding it to the PCR mixture alleviates the inhibition.
I just wanted to see if anyone else had first hand experience with this.
B) Also, for those with experience with melanin and PCR, would melanin inhibit bisulfite conversion?
I performed Methylation specific PCR and ofcourse see very weak bands (atleast I saw something compared to wild type PCR) in both methylated and unmethylated. I assume it is because melanin inhibits polymerase, but would the conversion also be impeded?
Edited by crom80, 23 October 2013 - 10:55 AM.