THE PRESENCE OF A DEFECTIVE LEU2 GENE ON 2-MU DNA RECOMBINANT PLASMIDS OF SACCHAROMYCES-CEREVISIAE IS RESPONSIBLE FOR CURING AND HIGH COPY NUMBER
THE PRESENCE OF A DEFECTIVE LEU2 GENE ON 2-MU DNA RECOMBINANT PLASMIDS OF SACCHAROMYCES-CEREVISIAE IS RESPONSIBLE FOR CURING AND HIGH COPY NUMBER
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DOI：
10.1128/jb.156.2.625-635.1983
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发表时间：
1983-01-01
影响因子：
3.2
通讯作者：
HOLLENBERG, CP
中科院分区：
文献类型：
--
作者：
ERHART, E;HOLLENBERG, CP
2 mu的拷贝数。CIR+酿酒葡萄球菌转化体中的dna衍生质粒由其选择标记决定，通常远低于内源质粒。在选择性条件下，只有含有pJDB219的leu2等位基因的质粒（称为leu2-d）的拷贝数高于内源的2.mu。DNA和置换产生了治愈的细胞。自发损失2亩。DNA的发生频率约为每代0.02%。固化质粒，如pMP78，拷贝数为35；非固化质粒（如pDB248或YEp6）的拷贝数为4到8。2.亩。测定菌株AH22和YNN27的DNA拷贝数分别为40和100。含LEU2 -d质粒的高拷贝数可以解释为其弱表达量不到野生型LEU2基因的5%。leu2-d等位基因在ATG起始密码子前缺失了从29个碱基对开始的5”端序列，但令人惊讶的是，其表达仍然受到调控。在含有染色体自主复制序列ARS1的YRp7上，有缺陷的leu2-d等位基因不能补足leu2宿主菌株。这表明含有ars1的质粒的复制控制比2.mu更为严格。含有质粒。
The copy number of 2.mu. DNA-derived plasmids in CIR+ S. cerevisiae tranformants is determined by its selective marker and is usually much lower than that of the endogenous plasmid. Only plasmids containing the leu2 allele of pJDB219, designated as leu2-d, under selective conditions displayed a higher copy number than did endogenous 2.mu. DNA and by displacement generated cured cells. Spontaneous loss of 2.mu. DNA occurred with a frequency of about 0.02% per generation. Curing plasmids, like pMP78, have copy numbers of 35; noncuring plasmids, like pDB248 or YEp6, have copy numbers of 4 to 8. The 2.mu. DNA copy number in strains AH22 and YNN27 were determined to be 40 and 100, respectively. The high copy number of leu2-d-containing plasmids can be explained by its weak expression of less than 5% that of the wild-type LEU2 gene. The leu2-d allele has a deletion of the 5''-end sequence starting from 29 base pairs before the ATG initiation codon, but surprisingly, its expression is still regulated. On YRp7, which contains the chromosomal autonomic replication sequence ARS1, the defective leu2-d allele could not complement a leu2 host strain. This suggests a more stringent control of replication of ARS1-containing plasmids than of 2.mu.-containing plasmids.