克雷伯氏菌产1,3-丙二醇菌株选育及甘油脱水酶基因转录分析  

Breeding of Klebsiella for Production of 1,3-propanediol and Transcription Analysis of Glycerol Dehydratase Gene

作　　者：聂玲燕[1,2] 方慧英[1,2] 诸葛斌[1,2] 宗红[1,2] 张成[1,2] 诸葛健[1,3,2] 

机构地区：[1]江南大学工业生物技术教育部重点实验室,无锡214122 [2]江南大学生物工程学院工业微生物研究中心,无锡214122 [3]江南大学食品科学与技术国家重点实验室,无锡214122

出　　处：《应用与环境生物学报》2013年第1期25-29,共5页Chinese Journal of Applied and Environmental Biology

基　　金：国家高技术研究发展计划("863"计划;20 09A A02Z210)资助~~

摘　　要：通过对野生克雷伯氏菌(Klebsiella)进行紫外诱变,获得一株耐高浓度甘油及高浓度1,3-丙二醇的突变菌株A-39-3,该突变株产1,3-丙二醇的量较野生菌株提高了58%,副产物2,3-丁二醇和乙醇的量分别降低了27%和19%.为探讨突变株高产1,3-丙二醇的原因,通过对突变菌株发酵过程中关键酶的酶活跟踪分析,发现1,3-丙二醇氧化还原酶(PDOR)的酶活力与野生菌株相差不大,甘油脱氢酶(GDH)的酶活力略有下降,而突变菌株的甘油脱水酶(GDHt)的酶活力明显高于野生菌株.并采用半定量RT-PCR方法,从转录水平上比较甘油脱水酶基因的差异,结果发现突变菌株A-39-3甘油脱水酶基因的相对mRNA转录水平是野生菌株K的2.58倍,分析说明该突变株1,3-丙二醇产量的提高与关键酶基因dhaB的mRNA转录水平关系密切.The ultraviolet mutagenesis breeding method was applied to wild Klebsiella （K） and a mutant strain A-39-3 which could endure high concentration of glycerol and 1,3-propanediol was obtained. Compared to the wild strain, the 1,3-propanediol concentrations of the mutant strain increased by 58%, while the by-products of 2,3-butanediol and ethanol concentrations decreased by 27% and 19% respectively. The tracking and analyzing of key enzyme activities during fermentation was therefore carried out. It was found that the activity of 1,3-propanediol oxidoreductase was no difference, and the activity of glycerol dehydrogenase decreased slightly, however, the activity of glycerol dehydratase significantly increased compared with the wild strain. In order to reveal the causes of high-yield 1,3-pronpanediol in A-39-3, the transcription level of glycerol dehydratase gene was analyzed by semi-quantitative RT-PCR. The results showed that the relative mRNA transcription level of glycerol dehydratase gene in A-39-3 was 2.58 times as much as the level in the wild strain. It suggested that the causes of the improvement of 1,3-propanediol in A-39-3 was closely related with the mRNA transcription level ofdhaB. Fig 5, Tab 3, Ref 16

关 键 词：克雷伯氏菌 1 3-丙二醇 紫外诱变 甘油脱水酶 半定量RT-PCR 

分 类 号：TQ923[轻工技术与工程—发酵工程] Q78[生物学—分子生物学]