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Bacteriophage ϕ1 as a gene-cloning vector in Bacillus subtilis

✍ Scribed by Kawamura, Fujio ;Saito, Hiuga ;Ikeda, Yonosuke

Publisher
Springer
Year
1980
Tongue
English
Weight
731 KB
Volume
180
Category
Article
ISSN
0026-8925

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✦ Synopsis

We attempted to use Bacillus subtilis phage phi 1 as a gene-cloning vector since the phi 1 genome was found to have few cleavage sites upon digestion with several kinds of restriction endonucleases. A phi 1 stock supplied by J. Ito (University of Arizona, Tucson, USA) consisted of two phages, phi 1E1 and phi 1E2, having one and two EcoRI-cleavage sites in their genomes respectively. From the latter isolate a deletion mutant phi 1E2 delta 1 was induced to increase the size range of DNA segments to be cloned. It was demonstrated, by in vitro recombination experiments with phage rho 11 DNA, that phi 1E2 delta 1 can be used for cloning EcoRI fragments of various sizes. We analyzed the DNAs of ten phi 1 clones isolated from independent transfectants and found that six of them carried rho 11 DNA fragments inserted at either of the two EcoRI-cleavage sites. Some of the hybrid phage DNAs were found to be cleaved with BamHI and HaeIII endonucleases and the rho 11 DNA portion, whereas the parental phi 1E2 delta 1 DNA was insensitive to any of these enzymes. These hybrid phages would therefore be useful vectors for cloning foreign DNA fragments generated by cleavage with BamHI or HaeIII endonucleases.

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