TRANSFORMATION OF ASPERGILLUS-NIDULANS BY USING A TRPC PLASMID
TRANSFORMATION OF ASPERGILLUS-NIDULANS BY USING A TRPC PLASMID
复制标题
DOI：
10.1073/pnas.81.5.1470
复制
发表时间：
1984-01-01
期刊：
影响因子：
--
通讯作者：
TIMBERLAKE, WE
中科院分区：
文献类型：
--
作者：
YELTON, MM;HAMER, JE;TIMBERLAKE, WE
构建了嵌合质粒，其携带来自子囊菌A的三功能trpC基因的完整拷贝。nidulans。该质粒命名为pHY 201，在大肠杆菌中复制，在大肠杆菌中它赋予对氨苄青霉素和氯霉素的抗性，并补充缺乏磷酸核糖基氨基苯甲酸异构酶活性的trpC突变体。用pHY 201转化A. nidulans trpC-菌株以每微克DNA> 20个稳定转化体的频率转化为trpC+。对转化子DNA的Southern杂交分析表明，pHY 201 DNA已整合到A.在大多数情况下，nidulans染色体。大多数整合事件似乎发生在受体菌株的trpC-等位基因位点。在几种情况下，pHY 201或其衍生物从A.通过限制性内切酶消化染色体DNA、连接和转化E.杆菌
A chimeric plasmid was constructed carrying a complete copy of the trifunctional trpC gene from the Ascomycete fungus A. nidulans. This plasmid, designated pHY201, replicates in Escherichia coli, where it confers resistance to ampicillin and chloramphenicol, and complements trpC mutants lacking phosphoribosylanthranilate isomerase activity. pHY201 was used to transform an A. nidulans trpC- strain to trpC+ at frequencies of > 20 stable transformants per .mu.g of DNA. Southern blot analysis of DNA from transformants showed that pHY201 DNA had integrated into the A. nidulans chromosomes in a majority of cases. Most of the integration events appeared to occur at the site of the trpC- allele of the recipient strain. In several instances, pHY201, or derivatives thereof, were recovered from A. nidulans transformants by restriction endonuclease digestion of chromosomal DNA, ligation, and transformation of E. coli.