摘要
应用聚合酶链式反应 (PCR)技术 ,从腐败梭菌强毒株C5 5_1中 ,扩增出大小为 132 7bp的腐败梭菌α毒素全基因 (csa132 7基因 ) ,并将其克隆入 pMD18_T载体中。转化至受体菌DH5α后 ,经Amp/IPTG/X_Gal选择培养 ,提取质粒 ,PCR和EcoRⅠ /PstⅠ双酶切鉴定 ,筛选阳性重组克隆。核苷酸序列分析证实 ,csa132 7基因开放阅读框架由 132 0bp组成 ,编码 4 4 0个氨基酸残基。与GenBank中登录的国外Fukushima 5株、Kagoshima 1株、Mie株和 4 4株的α毒素全基因相比 ,核苷酸同源率分别为 10 0 %、99 4 7%、99 2 5 %和 97 6 7% ,推导氨基酸的同源率分别达 10 0 %、10 0 %、99 7%和 97 0 6 %。表明本实验所克隆的csa132
The complete gene of α_toxin(cpa1327 gene) was amplified from Clostridium septidcum by polymerase chain reaction(PCR),and then was cloned into a clone vector pMD18_T.By PCR and EcoRⅠ/PstⅠ digestion,the gene was identified.After being sequenced,the cpa1327 gene was known to be consisting of 1 320 bp encoding 440 amino acid residues.Compared the α_toxin gene of C55_1 isolate with Fukushima 5, Kagoshima 1, Mie and 44 isolates abroad,it showed that they share 100 %, 99.47 %, 99.25 % ,(97.67 %) identity in nucleotide sequence and 100 %, 99.47 %, 99.25 % ,97.67 % identity in deduced amino acid sequence separately.It was concluded that the csa1327 gene we cloned were the α_toxin of Clostridium septidcum.
出处
《中国预防兽医学报》
CAS
CSCD
北大核心
2005年第2期112-115,共4页
Chinese Journal of Preventive Veterinary Medicine
关键词
腐败梭菌
Α毒素
分子克隆
核苷酸序列
Clostridium septicum
α-toxin
molecular cloning
sequencing
