作　　者： ; ; ; ; ;

机构地区： 中山大学生命科学学院水生经济动物研究所

出　　处： 《中山大学学报（自然科学版）》 2008年第2期89-92,共4页

摘　　要： 采用遗传工程方法,重组表达史氏鲟两种促性腺激素β亚基蛋白。选用原核表达载体pET-22b(+),分别插入史氏鲟GtHⅠ&Ⅱβ亚基成熟肽cDNA序列,构建成C端含有6个组氨酸(6-His)融合蛋白标签的表达质粒;分别转化大肠杆菌表达菌株BL21(DE3)并诱导表达2个基因。SDS-聚丙烯酰胺凝胶电泳显示重组融合蛋白相对分子质量分别为:GtHⅠβ亚基大约14 000,GtHⅡβ亚基15 000左右。分别用抗6个组氨酸融合标签的单克隆抗体及兔抗鲟鱼GTH多克隆抗体对2个表达蛋白进行Western-Blot分析,结果显示重组蛋白表达正确且有较高的免疫活性。GTHⅠβ重组蛋白在2 h就有明显的表达,6 h后随着时间增加表达量不再增加;25℃诱导重组蛋白产量比37℃诱导产量低。获得的重组蛋白质将可用于建立鲟鱼GtH的放射免疫测定方法。 Using the genetic engineering method, two prssed in mur sturg plasmids gonadotropin β - subunit of Acipenser schrenkii were ex- E. coli. Two cDNA Sequenes of gonadotropin （GTH） β-subunits mature peptide cDNA sequences of Amur sturgeon were inserted into pET -22b （ ＋ ） vector, using Nde I and Xho I restriction site. The recombinant of two GTH β-subunits were constructed as C-terminal histidine-tagged fusion protein. Transformants bar- boring expression vectors were induced in E. coli strain BL21 （ DE3 ）, the recombinant GtH Ⅰ -β and GTH Ⅱ-β were about 14 000 and 15 000 respectively . The results of SDS-PAGE and western-blotting with the McAb of 6- histidine showed that the recombinant protein had correct molecular weight, just as expected. They also showed high immunological activities through western-blotting with polyclonal antibody of purified sturgeon GTH. GtH Ⅰ β recombinant protein had obvious expression in 2 hours. The quantity of induced protein had no significant increase after 6 hours. The combinated protein will contribute to develop a RIA method for detection of Acipenser.

关 键 词： 史氏鲟 促性腺激素 亚基 蛋白质 遗传工程

领　　域： [生物学] [农业科学] [农业科学]