Heteroduplex analysis

Heteroduplex analysis (HDA) is a method in biochemistry used to detect point mutations in DNA (Deoxyribonucleic acid) since 1992.^[1] Heteroduplexes are dsDNA molecules that have one or more mismatched pairs, on the other hand homoduplexes are dsDNA which are perfectly paired.^[1][2] This method of analysis depend up on the fact that heteroduplexes shows reduced mobility relative to the homoduplex DNA.^[3] heteroduplexes are formed between different DNA alleles.^[4] In a mixture of wild-type and mutant amplified DNA, heteroduplexes are formed in mutant alleles and homoduplexes are formed in wild-type alleles.^[5] There are two types of heteroduplexes based on type and extent of mutation in the DNA. Small deletions or insertion create bulge-type heteroduplexes which is stable and is verified by electron microscope.^[6] Single base substitutions creates more unstable heteroduplexes called bubble-type heteroduplexes, because of low stability it is difficult to visualize in electron microscopy.^[5] HDA is widely used for rapid screening of mutation of the 3 bp p.F508del deletion in the CFTR gene.^[6]

Heteroduplexes and homoduplexes formed after amplification and re-annealing of wild type and mutant alleles