PCR Amplification Following Restriction to Detect Site-Specific DNA Methylation Academic Article uri icon

abstract

  • A procedure to test for DNA methylation at sites recognized by methylation-sensitive restriction endonucleases is described. The procedure is based on the assumption that the polymerase chain reaction (PCR) will amplify sequences between two primers only if the target DNA is intact after digestion. A carrot (Daucus carota) cell line that is heterozygous for two sequenced alleles of Dc8, a gene which is expressed during the later stages of embryogenesis provided an ideal source of DNA for developing and testing protocols. The promoters of the two alleles differs significantly in length between two sites used for primers, and only one promoter has a GATC (Sau 3A1 or Mbo I) site. This allowed development of a protocol where only the sequence lacking the GATC site was amplified to detectable levels following digestion of DNA with Mbo I which is insensitive to symmetric methylation withm4C orm5C. 1992 Kluwer Academic Publishers.

published proceedings

  • PLANT MOLECULAR BIOLOGY REPORTER

altmetric score

  • 6

author list (cited authors)

  • Chang, S., Magill, C. W., Magill, J. M., Fong, F., & Newton, R. J.

citation count

  • 1

complete list of authors

  • Chang, Shujun||Magill, Clint W||Magill, Jane M||Fong, Franklin||Newton, Ronald J

publication date

  • November 1992