Chemistry: molecular biology and microbiology – Enzyme – proenzyme; compositions thereof; process for... – Hydrolase
Patent
1994-09-06
1996-07-09
Patterson, Jr., Charles L.
Chemistry: molecular biology and microbiology
Enzyme , proenzyme; compositions thereof; process for...
Hydrolase
435193, 43525233, 536 232, C12N 922, C12N 1555, C12N 1570
Patent
active
055344283
ABSTRACT:
The present invention discloses the cloning and expression in Escherichia coli of the SstI/SacI restriction-modification system. The SstI methylase gene was cloned by methylase selection (the "Hungarian Trick" and the orientation and boundaries of the gene were established. The level of methylation of the host was improved significantly by expressing the SstI methylase from the strong RPSL promoter. Using the SstI methylase clone as a probe, a chromosomal map was generated in the area of the gene for the SstI methylase. DNA downstream from the methylase was cloned and found to contain the gene for the SstI endonuclease. The orientation of the SstI endonuclease was established and a strong promoter was placed close to the SstI endonuclease gene by generating nested deletions. By these methods, an E. coli strain was constructed which produced high levels of SstI endonuclease. The SacI locus in the S. achromogenes chromosome was found to be identical to the SstI locus in S. stanford in all measured respects.
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Harris Raymond D.
Longo Mary C.
Smith Michael D.
Life Technologies Inc.
Patterson Jr. Charles L.
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