His-猪生长激素融合基因在Bm-N细胞和蚕体内的表达与纯化

用构建的带有His—Tag pgh融合基因的重组杆状病毒Bm—BacPAK6—pgh研究了Bm—BacPAK6—pgh在家蚕细胞(Bm—N)和蚕体内的表达，并对蚕体表达产物进行了纯化．SDS—PAGE电泳分析显示，重组病毒Bin—BacPAK6—pgh在Bin—N细胞、蚕体中得到了融合表达，Western blot分析表明，Bm—N细胞、蚕体中表达的融合蛋白与大肠杆菌表达的猪生长激素具有相同的抗原性．Bm-BacPAK6-pgh在Bin—N细胞内的表达始于24h，而蚕体中的表达始于72h，二者的表达峰分别在96h和120h．经40％饱和度硫酸铵盐析和Ni—NTA Agarose亲和柱二步纯化可获得SDS—PA(正电泳纯的具有抗原性的重组猪生长激素融合蛋白。

Expression and purification of His-porcine growth hormone fusion gene in Bm-N cell and Bombyx mori

Using the recombinant baculovirus carrying the His-Tag pgh fusion gene, the expression of Bm-Bac-PAK6-pgh in BmN cell and in silkworm larvae have been studied, and the expression products in silkworm larvae have been purified. SDS-PAGE analysis indicated that the Bm-BacPAK6-pgh had been expressed in Bm-N cell and in silkworm larvae. Western blot analysis suggested that the fusion proteins expressed in Bm-N cell and in silkworm larvae have the same antigenicity with pGH expressed in E. coli. The expression of Bm-BacPAK6-pgh begins at 24h p. i. in Bm-N cell and 72h p. i. in silkworm larvae, and the expression peaks are 96h p. i. and 120h p. i. respectively. The recombinant pGH fusion proteins can be attained by two-step purification method, that was 40% ammonium sulphate salt out and Ni-NTA Agarose affinity chromatography respectively. The obtain fusion protein has the electrophoresis purity and also has immunogenicity.