Mutant strain of E. coli for detection of methyltransferase clon

Details Mutant strain of E. coli for detection of methyltransferase clon Mutant strain of E. coli for detection of methyltransferase clon

1992-10-06

1996-02-13

Fleisher, Mindy B.

Chemistry: molecular biology and microbiology

Measuring or testing process involving enzymes or...

Involving nucleic acid

4351723, 435193, 435199, 43525233, 4352528, C12Q 168, C12N 1509, C12N 1554, C12N 1555

Patent

active

054910609

ABSTRACT:
E. coli bacterial strains encoding a restriction gene that degrades methylated DNA, and prevents cloning of genes expressing the methyltransferase responsible for methylation, are mutated by a chemical or physical mutagen, so as to make the restriction enzyme temperature sensitive. Mutant cells are rendered competent, and plasmids expected or known to contain genes encoding methyltransferase enzymes are introduced. The transformants grow at the permissive temperature, where the restriction enzyme system is inactivated due to the mutated gene. Successful clones, expressing a methyltransferase, can be quickly identified by those which grow at the permissive temperature, but not at the non-permissive temperature. The valuable methyltransferases, as well as restriction enzymes associated therewith, can accordingly be recovered in large quantity.

REFERENCES:
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Lewin, Genes, John Wiley and Sons, New York, 2nd edition, (1985).
Slatco et al., Gene, 74, pp. 45-50 (1988).
Wu et al., Nucleic Acids Research, vol. 16, No. 2, pp. 703-717 (Jan. 25, 1988).
Hopwood et al., in Methods in Microbiology, vol. 3, pp. 363-433, Academic Press, London, (1970).
Miller, Expts. in Mol. Genetics, pp. 235-245, Cold Spring Harbor Laboratory, Cold Spring Harbor, N.Y. (1972).

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