瑞氏木霉木糖醇脱氢酶基因的分离与鉴定  

Isolation and Identification of Xylitol Dehydrogenase Gene from Trichoderma reesei

作　　者：汪天虹[1] 高培基[1] 王春卉[1] 钟玲 

机构地区：[1]山东大学微生物技术国家重点实验室

出　　处：《生物工程学报》1998年第3期320-325,共6页Chinese Journal of Biotechnology

摘　　要：将在木聚糖上生长的瑞氏木霉（Ｔｒｉｃｈｏｄｅｒｍａｒｅｓｅｉ）ＲｕｔＣ３０的ｃＤＮＡ文库全部质粒转化已携带有毕赤氏酵母（Ｐｉｃｈｉａｓｔｉｐｉｔｉｓ）木糖还原酶基因的重组酿酒酵母（Ｓａｃｈａｒｏｍｙｃｅｓｃｅｒｅｖｉｓｉａｅ）菌株Ｈ４７５，在Ｈ４７５中构建了瑞氏木霉的ｃＤＮＡ表达亚文库。在以木糖为唯一碳源的选择性酵母合成培养基上，从该亚文库中筛选到瑞氏木霉木糖醇脱氢酶ｃＤＮＡ基因，该基因片段长为１３ｋｂ。Ｓｏｕｔｈｅｒｎ、Ｎｏｒｔｈｅｒｎ印迹杂交分析和蛋白质凝胶电泳结果表明该基因确实来源于瑞氏木霉，所编码蛋白质分子量约为４０ｋＤａ。携带有毕赤氏酵母木糖还原酶和瑞氏木霉木糖醇脱氢酶基因的重组酵母能够在以木糖为唯一碳源的培养基上生长，并能将９０％以上的木糖转化为木糖醇、乙醇和其它副产品。A cDNA sub library from the fungus Trichoderma reesei grown on xylan was constructed in S.cerevisiae recombinant strain H475 harboring xylose reductase (XR)gene from Pichia stipitis .The sub library was screened for xylitol dehydrogenase (XDH) gene on SC selective medium in which xylose was used as a sole carbon source.The XDH gene,xdhl,was isolated from this sub pool and the length of xdhl is about 1.3kb.Southern,Northern and Western blot were carried out.The results indicated that xdhl has high affinity with T.reesei and the molecular weight of the xylitol dehydrogenase produced by S.cerevisiae recombinant strain HX1 is about 40 kDa.The strain HX1 harboring both xr from P.stipitis and xdhl from T.ressei was able to grown on xylose medium and converted more than 90% of the xylose into xylitol,ethanol and another by products.

关 键 词：瑞氏木霉 木糖醇脱氢酶 基因 酿酒酵母 分离 

分 类 号：Q933[生物学—微生物学]