摘要
运用生物信息学和PCR方法,鉴定了粘质沙雷氏菌乙偶姻生物合成途径中编码α-乙酰乳酸脱羧酶(α-ALDC)和α-乙酰乳酸合成酶(α-ALS)的基因(aceA和aceB)序列。经测序和序列分析显示aceA和aceB基因的开放阅读框(ORF)长度为780 bp和1 686 bp,编码259和561个氨基酸,编码蛋白分子量和等电点分别为28.96 kD和5.48,60.70 kD和5.88,属酸性蛋白。将两基因分别克隆至pET28a(+)表达载体中,并转化E.coli BL21(DE3)进行IPTG诱导表达,SDS-PAGE结果表明在大约29 kD和60 kD的位置出现特异性蛋白条带,表明aceA和aceB基因在E.coli BL21(DE3)中获得了高效的表达。
The aceA and aceB genes coding for α-acetolactate decarboxylase（α-ALDC） and α-acetolactate synthase（α-ALS） in the acetoin biosynthetic pathway from Serratia marcescens were identified and amplified through sequence alignment and Polymerase Chain Reaction（PCR）.The sequencing results showed that the ORF lengths of aceA and aceB genes were 780 bp and 1 686 bp,and encoded proteins of 259 and 561 residues respectively.Furthermore,their molecular weights and isoelectric points of 28.96 kD and 5.48,60.70 kD and 5.88 were predicted,which were acidic proteins judging from the calculated pI values.Then both genes were ligated with the expression plasmid of pET28a（＋） and transformed into E.coli BL21（DE3） for their induced expression with IPTG.SDS-PAGE analysis revealed that there were two clear induced protein bands with molecular weights of about 29 kD and 60 kD on expected position.These results indicated that both genes of aceA and aceB were efficiently expressed in E.coli BL21（DE3）.
出处
《江西农业大学学报》
CAS
CSCD
北大核心
2011年第5期987-992,共6页
Acta Agriculturae Universitatis Jiangxiensis
基金
国家"十一五"863专题(2006AA02Z243)
生物反应器国家重点实验室专项(2060204)
关键词
粘质沙雷氏菌
乙偶姻
克隆
序列分析
表达
Serratia marcescens
acetoin
cloning
sequence analysis
heterogenous expression
