Viral genome characterisation by the heteroduplex mobility and heteroduplex tracking assays.
The heteroduplex mobility assay (HMA) is a means of comparing two PCR amplicons or, in the variation known as the heteroduplex tracking assay (HTA), a means of estimating the quasispecies diversity of a viral genome. Heteroduplex assays have many applications including subtyping viral genomes, screening for low frequency variants in a population, scanning the relative genetic diversity across a genome and screening for recombinant clones. They can be used to detect dual infections, superinfections, contaminated blood products and laboratory contaminations. PCR amplicons of about 65% sequence similarity or greater will form heteroduplexes under appropriate conditions, and phylogenetic trees can be drawn from heteroduplex mobility data. While homoduplexes indicate more than 98% similarity between two DNA sequences, heteroduplexes indicate at least seven mismatches in a 500-bp amplicon, or a three-base pair gap in 1000-bp. Minority variants comprising 1% to 5% of the genome population can be detected and quantified by HTA. Thus far, heteroduplex assays have been described for HIV and other lentiviruses, hepatitis C and G viruses, Norwalk-like viruses, influenza, measles and poliovirus. They could be applied to a wide range of other viral species.
Barlow
K L
KL
Virus Reference Division, Central Public Health Laboratory, 61 Colindale Avenue, London NW9 5HT, UK.
Green
J
J
Clewley
J P
JP
D004279:DNA, Viral / Q000032:analysis / Q000235:genetics; D016679:Genome, Viral; D020180:Heteroduplex Analysis / Q000379:methods*; D006801:Humans; D009154:Mutation; D012327:RNA Virus Infections / Q000821:virology; D012328:RNA Viruses / Q000145:classification* / Q000235:genetics*; D012367:RNA, Viral
D004279:DNA, Viral; D012367:RNA, Viral