【摘要】 根据已发表的环状芽孢杆菌(Bacillus circulan)和枯草芽孢杆菌(Bacillus subtilis)木聚糖酶基因序列设计引物,首次扩增出短芽孢杆菌(Bacillus brevis)中的-β1,4-内切木聚糖酶(以下简称木聚糖酶,E.C.3.2.1.8)基因片段。序列分析表明,该基因与已登录的木聚糖酶基因AF490979.1和AF490980.1分别有97%和96%的同源性,与其它芽孢杆菌属的同源性也较高。将此基因片段插入表达载体pET-30a(+)构建重组质粒,转化大肠杆菌E.coliBL21(DE3)。重组基因工程菌破碎后进行SDS-PAGE电泳检测,结果表明,IPTG诱导后,木聚糖酶基因在大肠杆菌的胞内获得高效表达,且酶活力最高可达26.14 U/mL。重组木聚糖酶最适温度为50℃,最适pH为9.0。更多还原

【Abstract】 In this study,a pairs of primers were designed according to the published nucleotide sequences of putative xylanase genes of Bacillus circulan and Bacillus subtilis.With the specific primers,a target fragment of the β-1,4-endo-xylanase gene was firstly amplified from Bacillus brevi L8.Sequence analyses showed the homology of the cloned gene to AF490979.1and AF490980.1 were 97% and 96%,respectively.The recombinant expression plasmid was constructed and transformed into E.coli BL21(DE3).SDS-PAGE demonstrated that the β-1,4-endo-xylanase protein gene were expressed extracellularly in E.coli BL21(DE3) successfully.A maximum activity of 26.14 U/mL achieved gotten from cellular extract of pET-30a-xylB induced by IPTG.The optimum pH and temperature of the purified enzyme were 9.0 and 50 ℃,respectively.更多还原