( generate PCR products of different siz...)QIAGEN Multiplex PCR Handbook

generate PCR products of different sizes so that wild-type mice (wt) could be easily distinguished from heterozygote mutant mice (ht), or homozygote mutant mice (hm).

Analysis of microsatellites

Eukaryotic genomes contain repeated DNA sequences that differ both in the length and sequence of the repeat. These repeats, known as satellite DNA, are often highly polymorphic, with many different alleles present within a population. Alleles may also differ in the number of copies of a particular repeat sequence. Variation in satellite DNA sequences is important and can be used to determine genetic differences between organisms or closely related individuals. Satellite loci used for PCR analysis are usually short repeat units known as microsatellites. The length of the repeat unit is typically 26 nucleotides, and is referred to as a short tandem repeat (STR). In some cases, slightly longer repeats are used for analysis. These are referred to as variant number of tandem repeats (VNTRs) and can vary in both repeat length and number of repeats.

Amplification of microsatellite DNA may result in a low percentage of PCR products that are shorter than expected. Typically, the difference in size of these "stutter products" is exactly one repeat unit and these products are an intrinsic feature of the amplification of some satellite DNA sequences.

Taq DNA polymerases, including HotStarTaq in the QIAGEN Multiplex PCR Master Mix, add an extra A residue to the 3'-end of PCR products. When using the microsatellite cycling protocol, all PCR products should have this additional A residue at the 3' end. However, if larger amounts of PCR product are generated using an increased number of cycles, it
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