根据Genbank公布的来源于Bacillus deramificans的普鲁兰酶基因突变体序列(AX203843)合成普鲁兰酶成熟肽基因。将该基因插入芽孢杆菌分泌型表达载体pHY-WZX,重组质粒转化地衣芽孢杆菌B60608,重组地衣芽孢杆菌实现普鲁兰酶分泌表达。对重组菌产普鲁兰酶的条件进行优化,以含2%药媒和8%甘油的培养基最适合普鲁兰酶表达。

According to the sequence of pullulanase gene from Bacillus deramificans(NCBI accession number: AX203843),the gene encoding mature peptide of pulluanase was synthesized and designated pulA.The pulA was amplified by the method of PCR and cloned into the expression vector pHY-WZX,yielding hybrid plasmid pHY-WZX-pulA.Subsequently,pHY-WZX-pulA was introduced into Bacillus licheniformis B60608.Active pullulanase was expressed by recombiant B.licheniformis and secreted into medium.Culture condition of recombinant B.licheniformis were optimized for production of pullulanase.The optimized medium consists of 2 % cotton seed protein and 8 % of glycerol.