Heteroduplex analysis can increase the informativeness of PCR-amplified VNTR markers: application using a marker tightly linked to the COL2A1 gene.

Variable number of tandem repeat (VNTR) polymorphisms provide a high degree of informativeness in linkage studies. Whether performed by standard methods or by polymerase chain reaction (PCR), analysis of these markers involves assessment of the length of each allele. VNTR alleles usually differ in the number of tandem repeats. During PCR amplification of a VNTR closely linked to the type II collagen gene (COL2A1), we identified allelic microheterogeneity through the analysis of unique heteroduplexes between amplified strands of the two alleles. In one large pedigree, heteroduplex analysis identified six COL2A1 alleles; standard methods would have identified only three distinct alleles. The identification of these heteroduplexes allowed the determination of the COL2A1 inheritance pattern in the family, which otherwise would have been noninformative.