Chased PCR: A modified inverse PCR technique to characterize flanking regions of AT-rich DNA fragments |
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Authors: | Shams-Eldin Hosam Debierre-Grockiego Françoise Schwarz Ralph T |
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Affiliation: | Institute for Virology, Medical Center for Hygiene and Medical Microbiology, Philipps University, Marburg, Germany. shamseld@mailer.uni-marburg.de |
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Abstract: | 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. |
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