【摘要】 目的:在黑曲霉中表达猪磷脂酶A2基因(PLA2、PLA2M)。方法:分别构建含有自身信号肽或糖化酶信号肽的猪磷脂酶A2基因PLA2重组表达载体pSZHG-PLA2、pSZHGS-PLA2M,并通过农杆菌介导法转化黑曲霉。结果:经筛选各得到1株同源重组转化子。对筛选出的同源重组菌株进行发酵培养,并进行胞内以及胞外酶活检测,结果显示由自身信号肽引导分泌的同源重组菌株的胞内和胞外酶活分别为21.68U/mL和2.12 U/mL,由糖化酶信号肽引导分泌的同源重组菌株的胞内以及胞外酶活分别为15.07U/mL和3.10U/mL。结论:表明在黑曲霉表达系统中自身信号肽和糖化酶信号肽都不能引导重组猪磷脂酶A2的有效分泌。更多还原

【Abstract】 Objective:The purpose was to express porcine phospholipase gene(PLA2,PLA2M) in Aspergillus niger.Method:The study constructed porcine phospholipase A2 recombinant expression vector pSZHG-PLA2,pSZHGS-PLA2M,which contained its own signal peptide or glucoamylase signal peptide.pSZHG-PLA2 and pSZHGS-PLA2M were transformed into Aspergillus niger by Agrobacterrium-tumefacien mediated method.Result:After screened,respectively,both pSZHG-PLA2 and pSZHGS-PLA2M obtained a homologous recombination transformant.To screen out homologous recombination strains for fermentation,respectively,detected the intracellular enzyme activity and extracellular enzyme activity.The result showed that the intracellular and extracellular enzyme activity of homologous recombination strain,which guided by its signal peptide,were 21.68U/mL and 2.12 U/mL.The intracellular and extracellular enzyme activity of homologous recombination strain,which guided by glucoamylase signal peptide,were 15.07U/mL and 3.10 U/mL.Conclusion:The results showed that the study used of its signal peptide and glucoamylase signal peptide had failed to obtain effective secretion of the recombination porcine phospholipase A2 gene in Aspergillus niger.更多还原