【摘要】 目的　在大肠杆菌表达系统中高效表达生长抑素 (SS)基因。方法　以天然SS的氨基酸和基因序列为标准 ,将大肠杆菌使用频率较低的密码子分别更换成了使用频率较高的密码子 ,在SS基因的两端加上了合适的酶切位点 ,头部 :KpnI和NcoI;尾部 :PstI(这一酶切位点可与NsiI的切口互补 )。克隆至pThioHisA质粒的KpnI/PstI酶切位点后 ,再克隆至pThioHisA质粒的KpnI/NsiI酶切位点 ,使SS基因分别位于多克隆位点的尾部和前部。结果　重组质粒经酶切鉴定和基因序列测定证明 ,基因完全正确 ,在大肠杆菌TOP10中均得到较好的表达 ,IPTG诱导后 4h表达量基本达到最高 (37℃ ) ;在A60 0 0 .4～ 1.5之间用IPTG诱导均可获得到较好的表达 ,但以 0 .6左右为最佳诱导时机 ;在LB、TB和 2YT培养基中表达量依次为 :TB >LB >2YT ;表达的SS融合蛋白基本为水溶性的 ,具有良好的SS抗原性和免疫原性。结论　为基因工程SS大肠杆菌疫苗的研制奠定了基础更多还原

【Abstract】 Objective To highly express somatostatin(SS) gene in E.coli .Methods Replace the low frequency codons in natural SS gene with high frequency codons to E.coli ,and add the Kpn I and Nco I restriction sites at the N terminal,and Pst I restriction site,which can be annealed with Nsi I restriction site,at the C terminal.Clone the synthesized SS gene fragment to the Kpn I/Pst I restriction site of plasmid pThioHis A then to the Kpn I/Nsi I restriction site.Results Restriction and DNA sequencing showed correct sequence of the synthesized SS gene.The expression level of SS gene in E.coli TOP 10 reached peak value after being induced with IPTG at 37℃ for 4h.The SS fusion protein was expressed well when IPTG was added at an A value of 0.460 to 1.5,however,the optimum A value for expression was 0.6.The media,in the order of expression level,are LB,TB and 2YT.Almost all the expressed SS fusion proteins were water soluble and showed good antigenicity and immunogenicity.Conclusion The study laid a foundation of developing recombinant SS vaccine.更多还原