Heteroduplex Analysis

Heteroduplex analysis is used to discriminate between heterogeneous and homogeneous PCR products. Clonal PCR products will anneal to form homoduplexes after heat denaturation and rapid cooling whereas polyclonal PCR products will form heteroduplexes. The homoduplexes can be separated from the heteroduplexes on a nondenaturing polyacrylamide gel as they migrate more rapidly through the gel whereas the heteroduplexes from a background smear of slower-migrating species. In addition to the materials listed below, a vertical electrophoresis tank, glass plates, power pack transilluminator, and photoimager are necessary for heteroduplex analysis.

1. Accugel mix (29:1; National Diagnostics, Hessle, Yorkshire, UK).

2. Ammonium persulphate (AMP; Sigma, Poole, Dorset, UK).

5. Bromophenol blue (Sigma).

6. Ethidium bromide (Sigma).

7. pBR322 HAE III DNA ladder (Sigma).

8. 10X TBE (National Diagnostics).

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