A plasmid containing an excisable URA3 cartridge for gene disruption that permits repeated use of URA3 selection in constructing multiply disrupted yeast strains.
ATCC® Medium 1227: LB Medium (ATCC medium 1065) with 50 mcg/ml ampicillin
Biosafety Level
1
Biosafety classification is based on U.S. Public Health Service Guidelines, it is the responsibility of the customer to ensure that their facilities comply with biosafety regulations for their own country.
Comments
Restriction digests of the clone give the following sizes (kb): BamHI/BglII--5.0, 4.2; BamHI--9.2; EcoRI--9.2; PstI--6.6, 2.6; XbaI--uncut.
Used by inserting the excised cassette into a cloned target gene, isolating a restriction fragment containing the cassette with flanking target gene sequences, transforming Ura3- strains, and selecting for stable integrants.
Loss of the cassette (and one of the flanking IS10 repeats) by recombination can be selected by 5-fluoro-orotic acid resistance.
A plasmid containing an excisable URA3 cartridge for gene disruption that permits repeated use of URA3 selection in constructing multiply disrupted yeast strains.
The BglII/BamHI cassette contains URA3 flanked by 1.5 kb direct repeats from insertion sequence IS10.
Alani E, et al. A method for gene disruption that allows repeated use of URA3 selection in the construction of multiply disrupted yeast strains. Genetics 116: 541-545, 1987. PubMed: 3305158
Riles L, et al. Physical maps of the six smallest chromosomes of Saccharomyces cerevisiae at a resolution of 2.6-kilobase pairs. Genetics 134: 81-150, 1993. PubMed: 8514151
