Towards a Cre-based recombination system for generating integrated DNA repertoires site-specifically in yeast

A Cre-lox double recombination system is shown to work reasonably well in Saccharomyces cerevisiae. The system comprises an antibiotic resistance marker (the kanMX4 module) integrated at the chromosomal location to be targeted (leu2) of yeast strain FY10 and plasmid pRS315 carrying a foreign gene, h...

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Bibliographic Details
Published in:Biotechnology letters 2002-05, Vol.24 (9), p.727-733
Main Authors: DAVIES, Bryn G, ADAMS, Claire L, AIJAZ, Saima, COX, Jonathan P. L
Format: Article
Language:English
Subjects:
Antibiotic resistance
Biological and medical sciences
Biotechnology
Deoxyribonucleic acid
DNA
Fundamental and applied biological sciences. Psychology
Genes
Genetic engineering
Genetic technics
Genetics
Methods. Procedures. Technologies
Miscellaneous
Synthetic digonucleotides and genes. Sequencing
Yeasts
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Summary:A Cre-lox double recombination system is shown to work reasonably well in Saccharomyces cerevisiae. The system comprises an antibiotic resistance marker (the kanMX4 module) integrated at the chromosomal location to be targeted (leu2) of yeast strain FY10 and plasmid pRS315 carrying a foreign gene, human odorant receptor gene OR17-228, for targeting to that locus. Both OR17-228 and kanMX4 are flanked by loxP and loxP 511 sites. Cre recombinase expression was induced from plasmid pBS39. PCR analysis shows that a single copy of OR17-228 can be targeted to the leu2 locus in 22% of yeast cells after 24 h induction. DNA sequences of 5 successfully targeted loci show that OR17-228 is integrated in the intended orientation, and Southern blotting shows that there are no ectopic copies of OR17-228 present in these cases.[PUBLICATION ABSTRACT]
ISSN:0141-5492
1573-6776
DOI:10.1023/A:1015246420023