Amplification in Escherichia coli of enzymes involved in genetic recombination: construction of hybrid ColE1 plasmids carrying the structural gene for exonuclease I.
Endo-R-HindIII restriction endonuclease fragments obtained from F30 and pMB9 plasmid DNAs were ligated in vitro and used to transform a recB21 recC22 sbcB15 strain of E. coli K-12. The inability of this strain to stably maintain pMB9 alone permitted the isolation of transformants that carried hybrid plasmids containing the sbcB+ allele. These transformants became sensitive to ultraviolet light and recombination defieient and showed a 25-fold increase in the level of exonuclease I activity. The stability of the sbcB hybrid plasmids and their effects on exonuclease I activity have also been determined in wild-type and recA1 genetic backgrounds. The presence of the plasmids results in a 7-fold increase in the level of exonuclease I in a wild-type strain and 15-fold increase in a recA1 strain. The increased activity in the recA1 mutant appears to be a result of increased plasmid stability in this genetic background.
D001483:Base Sequence; D004269:DNA, Bacterial / Q000032:analysis; D004274:DNA, Recombinant; D004926:Escherichia coli / Q000201:enzymology*; D005092:Exonucleases / Q000378:metabolism; D005111:Extrachromosomal Inheritance; D005796:Genes; D005818:Genetic Engineering; D005838:Genotype; D010957:Plasmids; D011995:Recombination, Genetic
D004269:DNA, Bacterial; D004274:DNA, Recombinant; D005092:Exonucleases