I've recently been getting a non-specific het cluster when genotyping Bos taurus genomic DNA using several SNPs that had previously been showing good amplification with distinct WT, mutant and het clusters. My assay process has not changed and in fact I can get both the good result and the bad (het only) result using the same assay aliquots, so I believe the problem is with the DNA. Generally the sample size is 384, so I would expect to see genotypes other than het, especially because this population has previously shown different genotypes.
Does anyone know something that could be a contaminant in genomic DNA that could cause a loss of allele specificity in the assay while maintaining good amplification?
Is there a possibility that you happen to have run into a set of animals with higher than average twinning rate? Blood from animals that were in twin births will often give unclear genotyping results due to anastomoses. Otherwise your laboratory may have been contaminated with PCR product, do you take care to separate PCR setup from the amplification room?
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