Premium
Stabilisation in bacillus stearothermophilus of a recombinant plasmid carrying the homologous α‐amylase gene
Author(s) -
Aiba Shuichi,
Monden Yoshiaki,
Ohnishi Masatoshi,
Koizumi JunIchi,
Ming Zhang
Publication year - 1986
Publication title -
journal of chemical technology and biotechnology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.64
H-Index - 117
eISSN - 1097-4660
pISSN - 0268-2575
DOI - 10.1002/jctb.280361209
Subject(s) - recombinant dna , plasmid , microbiology and biotechnology , biology , amylase , gene , homology (biology) , dna , genetics , biochemistry , enzyme
The phenotypic stability of the recombinant plasmid pAT9 (11.5 MD), which contained the cloned α‐amylase gene from B. stearothermophilus , was studied in batch and continuous culture. Irrespective of the type of culture (batch or continuous; or using glucose or maltose as carbon source), deletion plasmids of the phenotype Km r Amy − the same size (7.3 MD) emerged with time. DNA sequencing analysis and Southern hybridization of a region adjacent to both ends of the pAT9 Hin dIII fragment containing the α‐amylase gene showed that almost the entire part of the Hin dIII fragment was lost. Homology between fragments contiguous to the pAT9 Hin dIII terminals allowed exclusion of a fragment that did not encode the α‐amylase gene and a new recombinant plasmid, pATHP9 (Km r Amy + ; 7.5 MD), was constructed which had enhanced phenotypic stability in B. stearothermophilus .