I am trying to standardize a protocol for ARMS PCR using primers designed by someone else.
The last time I ran the PCR products on the gel, all samples were seen as heterozygous (although one of them was a positive control and should have shown a band in only one of the lanes).
So I sat down and looked at the primer sequences to find out that the 1 bp difference between the primer pairs is for the last nucleotide of the primer. I think this is the reason why everything turns out heterozygous, irrespective of what it is. Is my conclusion correct???
Also, if I had to redesign these primers, what would be an appropriate placement of different nucleotide from the 3' end???
Thanks
