Decreasing amplification bias associated with multiple displacement amplification and short tandem repeat genotyping.

Although multiple displacement amplification (MDA) is being used increasingly to amplify genomes, the amplification bias generated by the varphi29 polymerase can be a concern with genotyping applications. It has been noted that the bias is pronounced with small template amounts, particularly with single nucleotide polymorphism (SNP) and short tandem repeat (STR) genotyping. Bias may occur between loci, or between alleles within a locus, and may differ between sample donors at the same loci. Previous research has suggested that omitting denaturation of the template prior to amplification can reduce the observed bias significantly. Comparison of the two methods (with and without denaturation) has found that nondenaturation of template reverses the direction of bias observed between allelic pairs following MDA. By combining two MDA reactions, one denatured and one nondenatured, the bias was found to be reduced significantly, aiding copy number analysis and subsequent genotyping.