ISSN:
0749-503X
Keywords:
gene disruption
;
fusion PCR
;
Saccharomyces cerevisiae
;
Life and Medical Sciences
;
Genetics
Source:
Wiley InterScience Backfile Collection 1832-2000
Topics:
Biology
Notes:
We adapted a fusion polymerase chain reaction (PCR) strategy to synthesize gene disruption alleles of any sequenced yeast gene of interest. The first step of the construction is to amplify sequences flanking the reading frame we want to disrupt and to amplify the selectable marker sequence. Then we fuse the upstream fragment to the marker sequence by fusion PCR, isolate this product and fuse it to the downstream sequence in a second fusion PCR reaction. The final PCR product can then be transformed directly into yeast. This method is rapid, relatively inexpensive, offers the freedom to choose from among a variety of selectable markers and allows one to construct precise disruptions of any sequenced open reading frame in Saccharomyces cerevisiae.
Additional Material:
2 Ill.
Type of Medium:
Electronic Resource
URL:
http://dx.doi.org/10.1002/yea.320111307
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