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Heteroduplex analysis

Heteroduplex analysis (HDA) is a method in biochemistry used to detect point mutations in DNA since 1992. Heteroduplexes are dsDNA molecules that have one or more mismatched pairs, on the other hand homoduplexes are dsDNA which are perfectly paired. This method of analysis depend up on the fact that heteroduplexes shows reduced mobility relative to the homoduplex DNA. heteroduplexes are formed between different DNA alleles. In a mixture of wild-type and mutant amplified DNA, heteroduplexes are formed in mutant alleles and homoduplexes are formed in wild-type alleles. 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. Single base substitutions creates more unstable heteroduplexes called bubble-type heteroduplexes, because of low stability it is difficult to visualize in electron microscopy. HDA is widely used for rapid screening of mutation of the 3 bp p.F508del deletion in the CFTR gene.

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

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.12 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

References

References

  1. J. Wallace, Andrew (2002). "SSCP/Heteroduplex Analysis". In D. M. Theophilus, Bimal; Rapley, Ralph (eds.). PCR Mutation Detection Protocols. Totowa, New Jersey: Humana Press, Totowa, New Jersey. pp. 151 - 164. ISBN 0896036170.
  2. Glavač, Damjan; Dean, Michael (1996), Pfeifer, Gerd P. (ed.), "Heteroduplex Analysis", Technologies for Detection of DNA Damage and Mutations, Springer US, pp. 241–251, doi:10.1007/978-1-4899-0301-3_18, ISBN 978-1-4899-0301-3{{citation}}: CS1 maint: work parameter with ISBN (link)
  3. El-Yazbi, Amira F.; Wong, Alysha; Loppnow, Glen R. (2017). "A luminescent probe of mismatched DNA hybridization: Location and number of mismatches". Analytica Chimica Acta. 994: 92–99. Bibcode:2017AcAC..994...92E. doi:10.1016/j.aca.2017.09.036. PMID 29126473.
  4. White, Marga Belle; Carvalho, Magda; Derse, David; O'Brien, Stephen J.; Dean, Michael (1992-02-01). "Detecting single base substitutions as heteroduplex polymorphisms". Genomics. 12 (2): 301–306. doi:10.1016/0888-7543(92)90377-5. ISSN 0888-7543. PMID 1740339.
  5. Menounos, Panayiotis G.; Patrinos, George P. (2010-01-01), Patrinos, George P.; Ansorge, Wilhelm J. (eds.), "Chapter 4 - Mutation Detection by Single Strand Conformation Polymorphism and Heteroduplex Analysis", Molecular Diagnostics (Second ed.), Academic Press, pp. 45–58, doi:10.1016/b978-0-12-374537-8.00004-3, ISBN 978-0-12-374537-8, retrieved 2019-12-02{{citation}}: CS1 maint: work parameter with ISBN (link)
  6. Wang, Y. H.; Barker, P.; Griffith, J. (1992-03-05). "Visualization of diagnostic heteroduplex DNAs from cystic fibrosis deletion heterozygotes provides an estimate of the kinking of DNA by bulged bases". Journal of Biological Chemistry. 267 (7): 4911–4915. doi:10.1016/S0021-9258(18)42917-1. ISSN 0021-9258. PMID 1537869.