黑曲霉葡萄糖氧化酶基因在毕赤酵母SMD1168中的表达

发布时间：2018-03-24 15:57

  本文选题：葡萄糖氧化酶　切入点：黑曲霉　出处：《食品与生物技术学报》2017年09期

【摘要】：在毕赤酵母SMD1168中利用乙醇氧化酶AOX1强启动子表达黑曲霉葡萄糖氧化酶(Glucose oxidase,GOD)。提取黑曲霉Aspergillus niger PCTC的基因组DNA,以此为模板进行PCR扩增获得葡萄糖氧化酶基因,将目的基因插入到具有AOX1强启动子的表达载体p PICZαA上,经电转化导入毕赤酵母SMD1168中。经zeocin抗性平板初筛、摇床复筛以及SDS-PAGE蛋白质电泳的检测,获得了一株产葡萄糖氧化酶活力的菌株,该株菌在30℃、200 r/min的培养条件下,经体积分数1.0%的甲醇诱导发酵1 d可获得1.12 U/mL的酶活。对该菌株进行了摇瓶产酶条件优化,其最佳发酵条件为:在pH 5、30℃下经体积分数1.5%甲醇诱导7 d,酶活为32 U/mL。
[Abstract]:In Pichia pastoris SMD1168, the strong promoter of ethanol oxidase AOX1 was used to express glucose oxidase gene of Aspergillus Niger. The genomic DNA of Aspergillus niger PCTC was extracted, and the glucose oxidase gene was obtained by PCR amplification. The target gene was inserted into the expression vector p PICZ 伪 A with strong promoter of AOX1, and was transformed into Pichia pastoris SMD1168 by electroporation. The screening of zeocin resistance plate, shaking bed screening and SDS-PAGE protein electrophoresis were performed. A strain producing glucose oxidase activity was obtained. The strain was cultured at 30 鈩，

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