新一代高产甘油重组菌的构建及其初步发酵

Construction and fermentation of a recombinant Candida glycerinogenes strain with high glycerol production

摘要:

产甘油假丝酵母(Candida glycerinogenes WL2002-5)是一株发酵生产甘油的工业化菌株。为进一步提高其产甘油能力, 本研究利用前期研究中成功克隆的产甘油假丝酵母中甘油合成关键酶3-磷酸甘油脱氢酶基因CgGPD1, 构建根癌农杆菌双元载体pCAM3300-zeocin-CgGPD1后, 电击转化根癌农杆菌LBA4404, 通过根癌农杆菌介导法(ATMT)转化产甘油假丝酵母, 构建了产甘油假丝酵母重组菌。并从中筛选出一株酶活力和产甘油性能较好的产甘油假丝酵母重组菌株C.g-G8。以葡萄糖为底物摇瓶发酵96 h后, 重组菌C.g-G8的甘油产量比野生型菌株Candida glycerinogene提高18.06%, 平均耗糖速率提高12.97%, 平均酶活力提高27.55%。本研究成功利用ATMT法转化产甘油假丝酵母构建新一代高产甘油菌株。

Abstract:

Candida glycerinogenes WL2002-5 (C. g) is an important industrial strain for glycerol production. To further improve glycerol production, we reconstructed a binary vector pCAM3300-zeocin-CgGPD1, introduced it to Agrobacterium tumefaciens LBA4404 by electroporation, and then transformed the T-DNA harboring the CgGPD1 to Candida glycerinogenes by Agrobacterium tumefaciens-mediated transformation (ATMT). After 96 h fermentation with glucose as the substrate, we screened a transformant named C.g-G8 with high glycerol production. Compared with the wild strain, the glucose consumption rate of C.g-G8 and the glycerol production were 12.97% and 18.06% higher, respectively. During the fermentation, the activity of glycerol-3-phosphate dehydrogenase of C.g-G8 was 27.55% higher than that of the wild strain. The recombinant Candida glycerinogenes with high glycerol production was successful constructed by ATMT method.