Single-strand conformation polymorphism analysis with high throughput modifications, and its use in mutation detection in familial hypercholesterolemia. International Federation of Clinical Chemistry Scientific Division: Committee on Molecular Biology Techniques

Clin Chem. 1997 Mar;43(3):427-35.

Abstract

The identification of the specific mutation causing an inherited disease in a patient is the framework for the development of a rationale for therapy and of DNA-based tests for screening relatives. We present here a review of the single-strand conformational polymorphism (SSCP) method, which allows DNA fragments that have been amplified with specific primers and PCR to be scanned rapidly for any sequence variation. The general principles of the method are described, as are the major factors that must be considered in developing an optimal SSCP strategy, namely the length of the PCR fragment and the temperature of the gel run. Options for sample denaturing gel characteristics and detection of DNA fragments are discussed. In addition, several modifications are presented that have been developed for high-throughput mutational analysis. The application of these techniques to screen for mutations in the LDL receptor gene in patients with familial hypercholesterolemia are described.

Publication types

  • Research Support, Non-U.S. Gov't
  • Review

MeSH terms

  • DNA / analysis
  • DNA Mutational Analysis / methods*
  • DNA Primers / chemistry
  • Electrophoresis, Polyacrylamide Gel / methods
  • Exons / genetics
  • Genetic Testing / methods
  • Humans
  • Hyperlipoproteinemia Type II / genetics*
  • Polymerase Chain Reaction
  • Polymorphism, Single-Stranded Conformational*
  • Receptors, LDL / genetics*

Substances

  • DNA Primers
  • Receptors, LDL
  • DNA