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Thursday, October 21, 2010

Analysis of Heteroduplex DNA


                         Heteroduplex DNA is formed by pairing an unknown nucleic acid strand with a reference strand of a known sequence. Single-stranded nucleic acids with complementary sequences come together and form a double-stranded hybrid molecule in which hydrogen bonds form only between complementary base pairs. 

                          The thermal stability of DNA is related to the number of hydrogen bonds between complementary base pairs; therefore, the greater the similarity in nucleotide sequences between two samples of DNA, the more hydrogen bonds will be present in the heteroduplexes produced. Heteroduplexes generated with a mismatch in the sequence between the two strands will have a different sequence-dependent electrophoretic mobility than homoduplex DNA generated with perfectly matched sequences. 

                           Heteroduplex analysis also can be done to detect different sequences in a mixture by using different probes labeled with different colors. Multiplex analysis has been applied to the simultaneous detection of six  heterozygous mutations in BRCA1 and BRCA2, screening of the three common prothrombotic polymorphisms pl(A), factor V Leiden, and MTHFR (C677T), rapid genetic screening of hemochromatosis, genetic diagnosis of factor V Leiden, clinical diagnosis of rifampin-resistant tuberculosis strains, and analysis of complex mutational spectra of human cells in culture. 

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