Functional analysis of promoter CpG methylation using a CpG-free luciferase reporter vector

Epigenetics. Jul-Sep 2006;1(3):127-30. doi: 10.4161/epi.1.3.3327. Epub 2006 Aug 28.

Abstract

Methylation of CpG dinucleotides within proximal promoters is often associated with transcriptional silencing. Methylation-dependent repression is well established for hypermethylated CpG island promoters that are characterized by a high density of CpG residues. The effect of CpG DNA methylation on CpG-poor promoters is less well characterized, probably due to the lack of convenient assay systems to test promoter activities in vitro. In this report, we describe a novel luciferase reporter vector, pCpGL, which completely lacks CpG dinucleotides and can be used to study the effect of promoter DNA methylation in transfection assays. Whereas a traditional reporter vector that contains a large number of backbone CpG residues significantly represses a CpG-free promoter when methylated, our new reporter vector is only repressed due to the presence of functionally important, methylated CpG residues. The pCpGL vector provides a useful tool to study the effects of CpG methylation on CpG-rich and CpG-poor promoters.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Base Sequence
  • Chromosomes, Human, X
  • DNA Methylation
  • Dinucleoside Phosphates / deficiency
  • Dinucleoside Phosphates / metabolism*
  • Genes, Reporter
  • Genetic Vectors*
  • Genomic Imprinting
  • Humans
  • Luciferases / genetics*
  • Molecular Sequence Data
  • Nucleic Acid Conformation
  • Promoter Regions, Genetic*
  • Restriction Mapping
  • Transfection

Substances

  • Dinucleoside Phosphates
  • cytidylyl-3'-5'-guanosine
  • Luciferases