BROAD HOST RANGE DNA CLONING SYSTEM FOR GRAM-NEGATIVE BACTERIA - CONSTRUCTION OF A GENE BANK OF RHIZOBIUM-MELILOTI
BROAD HOST RANGE DNA CLONING SYSTEM FOR GRAM-NEGATIVE BACTERIA - CONSTRUCTION OF A GENE BANK OF RHIZOBIUM-MELILOTI
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DOI：
10.1073/pnas.77.12.7347
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发表时间：
1980-01-01
期刊：
影响因子：
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通讯作者：
HELINSKI, DR
中科院分区：
文献类型：
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作者：
DITTA, G;STANFIELD, S;HELINSKI, DR
利用天然存在的抗生素抗性质粒RK2构建了一种广泛的宿主克隆载体，可以高频地转移到革兰氏阴性菌中。载体大小为20 k碱基对，编码四环素抗性，含有2个适合克隆的单限制性内切酶位点。动员是由连接ColE1复制子的RK2转移基因组成的辅助质粒实现的。利用该质粒载体，在大肠杆菌中构建并建立了野生型墨氏菌DNA基因库。其中一个杂交质粒含有约26个碱基对的DNA插入，与肺炎克雷伯菌的氮酶结构基因区域具有同源性。
A broad host range cloning vehicle that can be mobilized at high frequency into gram-negative bacteria was constructed from the naturally occurring antibiotic resistance plasmid RK2. The vehicle is 20 kbase pairs in size, encodes tetracycline resistance, and contains 2 single restriction enzyme sites suitable for cloning. Mobilization is effected by a helper plasmid consisting of the RK2 transfer genes linked to a ColE1 replicon. By use of this plasmid vehicle, a gene bank of the DNA from a wild-type strain of R. meliloti was constructed and established in Escherichia coli. One of the hybrid plasmids in the bank contains a DNA insert of approximately 26 kbase pairs which has homology to the nitrogenase structural gene region of Klebsiella pneumoniae.