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OALib Journal期刊
ISSN: 2333-9721
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利用甘油脱水酶基因构建产3-羟基丙醛工程菌及其表达比较
DOI: 10.3724/SP.J.1145.2013.00020, PP. 20-24
Keywords: 甘油脱水酶,3-羟基丙醛,大肠杆菌,tac启动子,发酵
Abstract:
3-羟基丙醛（3-hpa）是一种重要的化工产品，可由甘油经甘油脱水酶作用后生成.为获得产3-hpa基因工程菌，在已构建含甘油脱水酶基因及其激活因子大亚基质粒petac-dhab-gdra的基础上，构建了包含小亚基gdrb激活因子的重组质粒petac-dhab-gdra-gdrb.利用大肠杆菌通用tac启动子将该质粒在不同escherichiacolibl21、dh5a及jm109中进行表达.阳性转化子经iptg诱导后，提取总rna，以cdna为模板进行rt-pcr发现，目标基因在不同宿主都能较好转录.sds-page、酶活测定和3-hpa浓度测定结果表明，目标蛋白表达存在差异；酶活分别为4.7(±0.44)、3.5(±0.95)、8.1(±0.66)u/mg；发酵液中3-hpa的含量分别为0.012(±0.0044)、0.014(±0.003)、0.375(±0.018)gl-1，重组e.colijm109/petac-dhab-gdra-gdrb具有较好的甘油脱水酶基因表达和产3-hpa性能.该基因工程菌与克雷伯氏菌（klebsiellapneumoniae）相比，发酵副产物明显较少，有利于后期提取，为生产3-hpa提供了一条新思路.
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