Chased PCR: A modified inverse PCR technique to characterize flanking regions of AT-rich DNA fragments.
The chased polymerase chain reaction (PCR) technique described here is a convenient method enabling the characterization of flanking regions of a known A/T-rich DNA fragment in two different successive steps. The first step includes a modified inverse PCR with inverted tail-to-tail primers, each with 35 overhanging nucleotides for the insertion into a carrier plasmid. The second step consists of a technique similar to a site-directed mutagenesis. The chased PCR technique is simple, quick, versatile and efficient; it improves the inverse PCR technique and may be applied to any ligation-linker method. Consequently, the techniques for PCR-based gene isolation are more suitable for the isolation of missing sequences of A/T-rich or complex DNA.
Shams-Eldin
Hosam
H
Institute for Virology, Medical Center for Hygiene and Medical Microbiology, Philipps University, Marburg, Germany. shamseld@mailer.uni-marburg.de
Debierre-Grockiego
Françoise
F
Schwarz
Ralph T
RT
D000818:Animals; D001482:Base Composition; D001483:Base Sequence; D017931:DNA Primers / Q000235:genetics; D016054:DNA, Protozoan / Q000737:chemistry* / Q000235:genetics*; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D010957:Plasmids / Q000235:genetics; D010963:Plasmodium falciparum / Q000235:genetics*; D016133:Polymerase Chain Reaction / Q000379:methods*
D017931:DNA Primers; D016054:DNA, Protozoan