多年的生化、遗传学及分子分析研究所得的大量研究结果,使酵母成为生物制药研究者首选的模式生物系统和工具。因此,酵母成为普遍的基因表达研究和重组蛋白的常见载体。尽管研究用的酵母种属有多种,包括毕赤酵母属、汉逊酵母属和德巴利氏酵母属,但最常用的还是酵母菌属。
使用传统的酶分解法通常很难从细胞中提取完整的酵母mRNA和细胞内蛋白。裂解酶通常是粗制备,包含的RNA酶和蛋白酶不仅会破坏细胞壁,还会破坏目标分子。而且,酶裂解出原生质体通常需要加入去垢剂,而去垢剂会造成多种蛋白质失活。因此,机械裂解或破坏酵母细胞通常需要释放一些小分子。
1、破碎前的准备:
0.1g菌体(本实验以0.1g菌体为例);钢珠;研磨机(选用品牌上海净信科技Tissulyser-24);研磨罐。
2、研磨:
0.1g菌体加入30ML PBS溶液,混匀后用移液器均匀滴入液氮中,收集液氮冻住的小球,将小球放入上海净信科技预冷过的研磨罐中,调整频率,震荡时间。取下研磨罐再次放入液氮预冷取出来再次上机研磨
研磨现场图
Previous:全自动组织研磨机研磨猪扁脑体实验-上海净信
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