短小芽孢杆菌A—30耐碱性木聚糖酶基因的分子生物学研究

采用PCR方法对短小芽孢杆菌A－30菌株的耐碱性木聚糖酶基因进行克隆。在木聚糖选择平板上用刚果红染色法筛选出阳性克隆，提取阳性克隆的重组质粒进行酶切鉴定和测序。该基因在大肠杆菌中表达，过夜培养物胞外、胞内和周质空间的木聚糖酶酶活分别为0．159IUmL^-1、0.322IUmL^-1和0.007IUmL^-1。此木聚糖酶表现出较宽的pH作用范围，最适作用pH7左右，在pH9时仍有60％以上的酶活性。图4参14

STUDY ON ALKALI-TOLERANT XYLANASE GENE FROM BACILLUS P UMILUS A-30

The alkali-tolerant xylanase gene of Bacillus pumilus A-30 was cloned by PCR. The positive clones were screened on the selected LB agar plates supplemented with 0.2% oat spelt xylan by Congo red staining method. The recombinant plasmid from one positive clone was used for further characterization and DNA sequencing. In Eschericia coli, the xylanase gene could be expressed by the recombinant plasmid and the activities in extracellular, intracellular and periplasmic fractions were 0.159 IU mL -1, 0.322 IU mL -1 and 0.007 IUmL -1 respectively. The xylanase had optimal activity at pH 7.0, and retained more than 60% of the activity even at pH 9.0. Fig 4, Ref 14