CLONING AND EXPRESSION IN ESCHERICH1A COLI OF AN ALKALINE AND THERMOSTABLE EXOLIPASE FROM PSEVDOMONAS PSEUDOALCALIGENES
类产碱假单胞菌耐热碱性脂肪酶基因的克隆

A gene coding for an alkaline and thermostable exolipase of Pseudomonas pseudoalcaligenes was cloned into Escherichia coli LE392 by inserting Sau3 AI-generated DNA fragments into the BamHI site of pIJ285. Four colonies with esterase and lipase activities on the tributyrin agar plate were isolated by screening the constructed pseudomonas pseudoalcaligenes genomic library. Only one out of the four positive colonies showed lipase activity on the agar plate contatining olive oil and Rhodamine-B. Subclones of the 45 kb insert carrying lipase gene was obtained in E. coli HB101 using pUC118 as a vector, two of which (HB101 (pHZ1402)and HB101 (pHZ1403)), retained lipase activity, but the level seems to be different. They carried 3.0 kb and 2.9 kb inserts with a 2 kb overlapping sequence. The lipase activity of HB101 (pHZ1403) is about 4 times higher than that of HB101 (pHZ1402),as 5 times as that of original strain P. pseudoalcaligenes.