Sequence alterations can mask each other's presence during screening with SSCP or heteroduplex analysis: BRCA genes as examples

Biotechniques. 2000 Jul;29(1):94-8. doi: 10.2144/00291st04.

Abstract

For mutation detection, various screening techniques are widely used because DNA sequencing, the gold-standard method, is still considered to be expensive and laborious for high-throughput screening. Single-strand conformation polymorphism (SSCP) analysis, heteroduplex analysis (HA) and their variant techniques are popular and frequently used for this purpose. It is widely accepted that when searching for unknown sequence variations, any revealed distinct pattern should always be sequenced. We give examples here of the BRCA1 and BRCA2 genes where the SSCP/HA techniques can produce ambiguous predictions if used to detect known genetic variants compared to positive controls. Using direct DNA sequencing, we provide evidence that in such cases, mutations or polymorphisms can mask each other's presence. This phenomenon can often influence the results of any DNA testing because genetic variations such as single-nucleotide polymorphisms occur frequently in the human genome. We suggest that even in the case of known electrophoretic patterns of well-characterized genetic alterations, every sequence alteration should be confirmed by direct DNA sequencing, especially if genetic testing is carried out for diagnostic purposes.

Publication types

  • Research Support, U.S. Gov't, P.H.S.
  • Technical Report

MeSH terms

  • BRCA2 Protein
  • DNA / blood
  • Exons
  • Genes, BRCA1*
  • Humans
  • Introns
  • Lymphocytes / chemistry
  • Neoplasm Proteins / genetics*
  • Nucleic Acid Heteroduplexes* / analysis
  • Polymerase Chain Reaction
  • Polymorphism, Single-Stranded Conformational*
  • Sequence Analysis, DNA
  • Transcription Factors / genetics*

Substances

  • BRCA2 Protein
  • Neoplasm Proteins
  • Nucleic Acid Heteroduplexes
  • Transcription Factors
  • DNA