链霉菌Streptomyces olivaceoviridis A1木聚糖酶基因xynA在大肠杆菌及毕赤酵母中的高效表达  

Expression of Xylanase Gene xynA from Streptomyces olivaceoviridis A1 in Escherichia coli and Pichia pastoris

作　　者：张红莲[1] 姚斌[1] 王亚茹[1] 袁铁峥[1] 张王照[1] 伍宁丰[2] 范云六[2] 

机构地区：[1]中国农业科学院饲料研究所,北京100081 [2]中国农业科学院生物技术研究所,北京100081

出　　处：《生物工程学报》2003年第1期41-45,共5页Chinese Journal of Biotechnology

基　　金：国家高技术研究与发展计划 (863计划 )项目资助 (No .2 0 0 1AA2 14 0 41)~~

摘　　要：从橄榄绿链霉菌StreptomycesolivaceoviridisA1中克隆出木聚糖酶基因xynA ,将带与不带原基因信号肽编码序列的xynA分别以正确的阅读框架克隆到大肠杆菌表达载体pET 2 2b(+)上的pellB信号肽编码序列之后 ,得到 2种构建的重组载体 ,在重组大肠杆菌中木聚糖酶得到了表达 ,表达产物具有生物活性。进一步将不带原基因信号肽编码序列的xynA插入到毕赤酵母转移载体pPIC9中 ,转化毕赤酵母得到重组子 ,在重组子中木聚糖酶基因得到了高效分泌表达 ,在摇床培养水平上的表达量达到 2 0 0mg L ,且表达产物具有生物学活性。The gene xynA encoding xylanase was cloned from Streptomyces olivaceoviridis A1. The xynA with and without origin signal peptide sequence were fused behind pel B signal peptide in the plasmid pET 22b(+)respectively, then transfered into the host E.coli. The xylanase expressed in E.coli had normal bioactivity. Further, the xynA without origin signal peptide sequence was cloned into the plasmid pPIC9 under the control of AOX1 promoter and introduced into the host Pichia pastoris by electroporation. The results of SDS PAGE and activity assay of the xylanase expressed by recombinant P.pastoris showed that the xynA had been overexpressed and secreted, and the xylanase expressed had normal bioactivity. The expression level of xylanase in recombinant P.pastoris exceeded 0.2mg/mL in shake culture.

关 键 词：链霉素 木聚糖酶基因 xynA 大肠杆菌 毕赤酵母 高效表达 

分 类 号：Q786[生物学—分子生物学]