斜带石斑鱼生长激素基因重组蛋白的发酵

Fermentation of Recombinant Protein of Growth Hormone Gene in Orange-spotted Grouper

将含有石斑鱼生长激素基因的毕赤酵母X33进行震荡培养后,装入≥50L的自控罐中,28~30.5℃、pH 5.5~6.0、溶解氧≥30mg/L条件下发酵84h,生产石斑鱼生长激素基因重组蛋白。发酵期间,通过甲醇诱导毕赤酵母获得目的蛋白的高效表达,最后通过Western Blot和SDS-PAGE聚丙酰胺凝胶电泳对目的蛋白进行定性定量检测。最终获得目的蛋白表达量约为1.5~2.0g/L,约占胞外可溶性总蛋白的16.4%。本试验用自控罐发酵生产含有斜带石斑鱼生长激素基因重组蛋白的毕赤酵母生产工艺,为实现工业化生产鱼类生长激素饵料添加剂奠定基础。

Yeast X33 Pichia pastoris were applied to express recombinant of growth hormone in orange- spotted grouper Epinephelus coioides cultivated in shaking flask first, and than fermented in 50 liters or bigger auto-control fermentator under conditions of temperature from 28℃ to 30.5 ℃ ,and pH 5.5-6.0 for 76 hours. During the fermentation period, the Yeast X33 were induced to express high amount protein by methanol, and the targets were detected qualitatively and quantitatively by Western Blot and SDS- PAGE. The concentration of target protein was found to be changed from 1.5 g/L to 2.0 g/L, accounting for 16.4% of total soluble protein in vitro. The fementation technology which producted recombinant or- ange-spotted grouper growth hormone in yeast X33 P. pastoris by an automatic control fermentor lays the foundation of industrial production of fish growth hormone feed additives.