宇佐美曲霉木聚糖酶基因在大肠杆菌中的表达

在已知宇佐美曲霉E001菌株木聚糖酶Xyn II cDNA序列（Genbank登录号为DQ114485）的基础上，合成1对引物（含 EcoR I和Sal I酶切位点），以重组质粒pUCm-T-xyn II为模板，扩增得到编码Xyn II成熟肽的cDNA片段（555 bp）。将其与表达质粒pET-28a连接，构建了重组表达质粒pET-28a-xyn II，转化E. coli BL21-CodonPlus(DE3)-RIL，获得重组工程菌B21/xyn II。经IPTG诱导表达，木聚糖酶的比酶活力最高可达35.6 U/mg。最后采用金属Ni2+螯和层析柱对所表达的木聚糖酶进行纯化，达到了电泳纯。重组表达的木聚糖酶最适温度为45℃，最适pH值为4.6。

Expression of Aspergillus usamii Xylanase Gene (xyn II) in Escherichia coli

Having known the Xyn II cDNA sequence (Genbank accession number DQ114485),a pair of primers (with the EcoR I and Sal I sites) were designed.With the recombinant plasmid pUCm-T-xyn II as a template,Xyn II cDNA fragment (555 bp) encoding mature peptide was then amplified,inserted into the plasmid pET-28a.The resultant recombinant plasmid pET-28a- xyn II was transformed into E. coli BL21- CodonPlus(DE3)-RIL,and finally the recombinant strain B21/xyn II was obtained.Induced by IPTG,a maximum activity of 35.6 U/mg was obtained from cellular extract of E. coli B21/xyn II.The recombinant protein was purified by immobilizing metal affinity chromatography.The optimum temperature and pH for recombinant enzyme were 45℃ and 4.6,respectively.