外源功能基因在木质纤维素依赖型乳酸菌Pediococcus acidilactici DQ2中的表达

Expression of Functional Genes in Lignocellulose-dependent Lactic Acid Bacterium Pediococcus acidilactici DQ2

乳酸片球菌Pediococcus acidilactici DQ2是一株耐高温和耐木质纤维素降解抑制物的乳酸高产菌株.为了在P.acidilactici DQ2中建立外源蛋白表达系统,对常用的乳酸菌表达载体pMG36e进行改造,将其启动子P32更换为来源于P.acidilactici DQ2的L-乳酸脱氢酶基因的启动子PldhL.并通过新载体pTY36e成功表达了两种不同的外源基因:来源于维多利亚多管发光水母(Aequorea victoria)的绿色荧光蛋白基因gfp和来源于多粘芽孢杆菌(Bacillus polymyxa 1.794)的β-葡萄糖苷酶基因bglA,其中所表达的胞内β-葡萄糖苷酶活力为4.48 U g-1(细胞干重).此系统的成功构建为后续对该菌的基因工程改造奠定了基础.

Pediococcus acidilactici DQ2, isolated by our lab, is thermotolerant and highly resistant to the inhibitors derived from lignocellulose, and can produce high titer of lactic acid. To construct the expression system of this strain, we modified the lactic acid bacterial expression vector pMG36e by replacing the promoter P32 with PldhL derived from P. acidilactici DQ2. Two different heterologous genes were expressed successfully with the new plasmid pTY36e： green fluorescent protein gene （gfp） from Aequorea victoria and β-glucosidase gene （bglA） from Bacillus polymyxa 1.794. The results showed β-glucosidase enzyme activity of the recombinant bacteria as 4.48 U g-1 （dry cells） detected in the intracellular fraction. The successful construction of this system lays the foundation for subsequent genetically engineering of P. acidilactici DQ2. Fig 4, Tab 3, Ref 25