Protocol

Cytochrome P450 Protocols

Volume 320 of the series Methods in Molecular Biology pp 193-207

Genotyping for Cytochrome P450 Polymorphisms

  • Ann K. DalyAffiliated withSchool of Clinical and Laboratory Sciences, University of Newcastle upon Tyne Medical School
  • , Barry P. KingAffiliated withSchool of Clinical and Laboratory Sciences, University of Newcastle upon Tyne Medical School
  • , Julian B. S. LeathartAffiliated withSchool of Clinical and Laboratory Sciences, University of Newcastle upon Tyne Medical School

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Abstract

Protocols for the extraction of DNA from human blood and for genotyping for a number of common cytochrome P450 polymorphisms using either polymerase chain reaction (PCR)-restriction fragment length polymorphism or PCR-single-strand conformational polymorphism (SSCP) analysis are described. Rapid high-throughput techniques are also available for analyses of this type, but they require access to specialized equipment and are not considered here. General guidelines for performing amplification using PCR are described together with electrophoresis protocols for analysis of restriction digests of PCR products with agarose and polyacrylamide gels including the use of polyacrylamide-based gels for SSCP analysis. Protocols for the following specific isoforms and alleles are also provided: CYP1A1 (*2B and (*4 alleles), CYP2C8 ((*3 and (*4 alleles), CYP2C9 ((*2, (*3, and (*11 alleles), CYP2C19 ((*2 and (*3 alleles), CYP2D6 ((*3, (*4, (*5, and (*6 alleles), CYP2E1 ((*5A, (*5B, and (*6 alleles), and CYP3A5 ((*3 allele).

Key Words

Polymerase chain reaction restriction digest polymorphism CYP1A1 CYP2C8 CYP2C9 CYP2C19 CYP2D6 CYP2E1 CYP3A5