SiteFinding-PCR: a simple and efficient PCR method for chromosome walking

Nucleic Acids Res. 2005 Aug 2;33(13):e122. doi: 10.1093/nar/gni124.

Abstract

In this paper, we present a novel PCR method, termed SiteFinding-PCR, for gene or chromosome walking. The PCR was primed by a SiteFinder at a low temperature, and then the target molecules were amplified exponentially with gene-specific and SiteFinder primers, and screened out by another gene-specific primer and a vector primer. However, non-target molecules could not be amplified exponentially owing to the suppression effect of stem-loop structure and could not be screened out. This simple method proved to be efficient, reliable, inexpensive and time-saving, and may be suitable for the molecules for which gene-specific primers are available. More importantly, large DNA fragments can be obtained easily using this method. To demonstrate the feasibility and efficiency of SiteFinding-PCR, we employed this method to do chromosome walking and obtained 16 positive results from 17 samples.

Publication types

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

MeSH terms

  • Arabidopsis / genetics
  • Chromosome Walking / methods*
  • Cyanobacteria / virology
  • DNA Primers / chemistry
  • DNA, Bacterial / genetics
  • Mutagenesis, Insertional
  • Myoviridae / genetics
  • Polymerase Chain Reaction / methods*
  • Rhizobium / genetics

Substances

  • DNA Primers
  • DNA, Bacterial