CLONING OF THE C-URA3 GENE AND CONSTRUCTION OF A TRIPLE AUXOTROPH (HIS5, ADE1, URA3) AS A USEFUL HOST FOR THE GENETIC-ENGINEERING OF CANDIDA-MALTOSA

被引：27

作者：

OHKUMA, M ^{[1
]}

MURAOKA, S ^{[1
]}

HWANG, CW ^{[1
]}

OHTA, A ^{[1
]}

TAKAGI, M ^{[1
]}

机构：

[1] UNIV TOKYO,DEPT AGR CHEM,TOKYO 113,JAPAN

来源：

CURRENT GENETICS | 1993年 / 23卷 / 03期

关键词：

CANDIDA-MALTOSA; C-URA3; TRIPLE AUXOTROPH; GENE DISRUPTION;

D O I：

10.1007/BF00351497

中图分类号：

Q3 [遗传学];

学科分类号：

071007 ; 090102 ;

摘要：

The C-URA3 gene of the n-alkane assimilating-yeast Candida maltosa was cloned by complementation of the ura3 mutation of Saccharomyces cerevisiae. The nucleotide sequence of C-URA3 and its deduced amino-acid sequence showed significant homology to those of the orotidine 5'-phosphate decarboxylases of other fungal species. To construct a useful host for genetic engineering of C. maltosa using C-URA3 as a marker, one allele of C-URA3 in a double auxotroph (his5, ade1) was disrupted by C-ADE1, and subsequently two kinds of ura3 mutants were isolated by selecting for spontaneous 5-fluoro-orotic acid (5FOA) resistance. One of the mutants was homozygous for the disruption (ura3::C-ADE1/ura3::C-ADE1); the other was heterozygous (ura3::C-ADE1/ura3). The ura3::C-ADE1 allele in the latter strain was re-substituted by C- URA 3 to rescue the adenine auxotroph (his5, ade1, C-URA3/ura3). Finally, by selecting a 5FOA-resistant mutant, a triple auxotroph (his5, ade1, ura3/ura3) was isolated.

引用

页码：205 / 210

页数：6

共 20 条

[1] A METHOD FOR GENE DISRUPTION THAT ALLOWS REPEATED USE OF URA3 SELECTION IN THE CONSTRUCTION OF MULTIPLY DISRUPTED YEAST STRAINS [J].