ISSN:
0749-503X
Keywords:
yeast functional analysis
;
gene disruption
;
gap-repair cloning
;
selection gene ‘pop-out’
;
I-SceI
;
Life Sciences
;
Life Sciences (general)
Source:
Wiley InterScience Backfile Collection 1832-2000
Topics:
Biology
Notes:
New tools are needed for speedy and systematic study of the numerous genes revealed by the sequence of the yeast genome. We have developed a novel transformation strategy, based on ‘split-marker’ recombination, which allows generation of chromosomal deletions and direct gene cloning. For this purpose, pairs of yeast vectors have been constructed which offer a number of advantages for large-scale applications such as one-step cloning of target sequence homologs and combinatorial use. Gene deletions or gap-repair clonings are obtained by cotransformation of yeast by a pair of recombinant plasmids. Gap-repair vectors are based on the URA3 marker. Deletion vectors include the URA3, LYS2 and kanMX selection markers flanked by I-SceI sites, which allow their subsequent elimination from the transformant without the need for counter-selection. The application of the ‘split-marker’ vectors to the analysis of a few open reading frames of chromosome XI is described.
Additional Material:
7 Ill.
Type of Medium:
Electronic Resource
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