Matsuzaki H, Nakajima R, Nishiyama J, Araki H, Oshima Y
Department of Fermentation Technology, Faculty of Engineering, Osaka University, Japan.
J Bacteriol. 1990 Feb;172(2):610-8. doi: 10.1128/jb.172.2.610-618.1990.
We have developed an effective method to delete or invert a chromosomal segment and to create reciprocal recombination between two nonhomologous chromosomes in Saccharomyces cerevisiae, using the site-specific recombination system of pSR1, a circular cryptic DNA plasmid resembling 2 microns DNA of S. cerevisiae but originating from another yeast, Zygosaccharomyces rouxii. A 2.1-kilobase-pair DNA fragment bearing the specific recombination site on the inverted repeats of pSR1 was inserted at target sites on a single or two different chromosomes of S. cerevisiae by using integrative vectors. The cells were then transformed with a plasmid bearing the R gene of pSR1, which encodes the site-specific recombination enzyme and is placed downstream of the GAL1 promoter. When the transformants were cultivated in galactose medium, the recombination enzyme produced by expression of the R gene created the modified chromosome(s) by recombination between two specific recombination sites inserted on the chromosome(s).
我们开发了一种有效的方法，利用pSR1的位点特异性重组系统，在酿酒酵母中删除或反转染色体片段，并在两条非同源染色体之间产生相互重组。pSR1是一种环状隐蔽DNA质粒，类似于酿酒酵母的2μm DNA，但起源于另一种酵母——鲁氏接合酵母。通过整合载体，将携带pSR1反向重复序列上特异性重组位点的2.1千碱基对DNA片段插入到酿酒酵母单条或两条不同染色体的靶位点。然后用携带pSR1的R基因的质粒转化细胞，该基因编码位点特异性重组酶，位于GAL1启动子下游。当转化体在半乳糖培养基中培养时，R基因表达产生的重组酶通过染色体上插入的两个特异性重组位点之间的重组产生修饰的染色体。
