COLONY BANK CONTAINING SYNTHETIC COL EL HYBRID PLASMIDS REPRESENTATIVE OF ENTIRE ESCHERICHIA-COLI GENOME
COLONY BANK CONTAINING SYNTHETIC COL EL HYBRID PLASMIDS REPRESENTATIVE OF ENTIRE ESCHERICHIA-COLI GENOME
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DOI：
10.1016/0092-8674(76)90055-6
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发表时间：
1976-01-01
期刊：
影响因子：
64.5
通讯作者：
CARBON, J
中科院分区：
文献类型：
--
作者：
CLARKE, L;CARBON, J
Using the poly(dA.cntdot.dT) connector method, a population of annealed hybrid circular DNA was constructed in vitro; each hybrid DNA circle contained 1 molecule of poly(dT)-tailed Col E1-DNA annealed to any one of a collection of poly(dA)-tailed linear DNA fragments, produced originally by shearing total Escherichia coli DNA to an average size of 8.5 .times. 106 daltons. This annealed DNA preparation (12 .mu.g) was used to transform an F+ recA E. coli strain (JA200), selecting transformants by their resistance to colicin E1. A collection or bank of > 2000 colicin-E1 resistant clones was thus obtained, 70% of which were shown to contain hybrid Col E1-DNA E. coli plasmids. This colony bank is large enough to include hybrid plasmids representative of the entire E. coli genome. Individual plasmids were readily identified by replica mating the collection onto plates seeded with cultures of various F- auxotrophic recipients, selecting for complementation of the auxotrophic markers by F-mediated transfer of hybrid plasmids to the F- recipients. Thus, > 80 hybrid Col E1-DNA E. coli plasmid-bearing clones were identified in the colony bank, and .apprx. 40 known E. coli genes were tentatively assigned to these various plasmids. The hybrid plasmids are transferred efficiently from F+ donors to appropriate F- recipients. The use of this method to establish similar colony banks in E. coli containing hybrid plasmids representative of various simple eukaryotic [yeast and drosophila] genomes is discussed.
