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PCR-mediated direct gene disruption in Schizosaccharomyces pombe.

DIR@IMTECH: CSIR-Institute of Microbial Technology

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Title PCR-mediated direct gene disruption in Schizosaccharomyces pombe.
 
Creator Kaur, R
Ingavale, S S
Bachhawat, Anand K
 
Subject QR Microbiology
 
Description We have examined the feasibility and efficiency of PCR-mediated direct gene disruptions in the fission yeast Schizosaccharomyces pombe. In the present study, the S.pombe ura4+ gene was amplified by PCR with oligonucleotides that had short flanking regions ( approximately 40 bp) to the target gene. Using this purified PCR product we were able to disrupt genes in an S. pombe strain bearing aura4 deletion, with an efficiency ranging between 1 and 3% among selected transformants. The results indicated that despite S.pombe's preference for non-homologous or illegitimate recombination, even very short stretches of homologous regions could be used to target genes at a defined frequency in this organism. The successful disruption of four independent genes (sts1+, gcs1+, gsh2+and hmt1+) by this method further demonstrates that, despite the relatively low efficiency, the method is very feasible, and it's simplicity, especially when coupled to phenotype-based screening, should greatly facilitate disruption of genes in S.pombe.
 
Publisher Oxford University Press
 
Date 1997-03-01
 
Type Article
PeerReviewed
 
Format application/pdf
 
Identifier http://crdd.osdd.net/open/370/1/bachhawat97.pdf
Kaur, R and Ingavale, S S and Bachhawat, Anand K (1997) PCR-mediated direct gene disruption in Schizosaccharomyces pombe. Nucleic acids research, 25 (5). pp. 1080-1. ISSN 0305-1048
 
Relation http://nar.oxfordjournals.org/content/25/5/1080.full.pdf+html
http://crdd.osdd.net/open/370/