Goodridge A, Civelli O, Yip C, Scherrer K
Eur J Biochem. 1979 May 2;96(1):1-8. doi: 10.1111/j.1432-1033.1979.tb13006.x.
Undegraded, biologically-active, polyadenylated RNA was isolated from chicken liver by a rapid, simple procedure. Liver cells were dispersed mechanically and then broken gently by controlled Dounce homogenization in the absence of detergent or ribonuclease inhibitors. After removing lysosomes and mitochondria by centrifugation, RNA was precipitated at pH 5.2. Polyadenylated mRNA was isolated directly from the detergent-solubilized precipitate by oligo(dT)-cellulose chromatography. The resulting RNA was translated into liver-specific peptides in both the wheat germ lysate and Xenopus laevis oocytes. Translatable albumin mRNA was detected in the liver cytoplasm of both fed 3-week-old chicks and unfed, day-old chicks. Translatable malic enzyme mRNA was only detected in the livers from the fed chicks.
通过一种快速、简单的方法从鸡肝脏中分离出未降解的、具有生物活性的多聚腺苷酸化RNA。肝细胞通过机械分散,然后在不存在去污剂或核糖核酸酶抑制剂的情况下,通过可控的Dounce匀浆法轻轻破碎。通过离心去除溶酶体和线粒体后,在pH 5.2条件下沉淀RNA。通过寡聚(dT)-纤维素色谱法直接从去污剂溶解的沉淀物中分离多聚腺苷酸化的mRNA。所得RNA在小麦胚芽裂解物和非洲爪蟾卵母细胞中均被翻译成肝脏特异性肽。在3周龄喂食的雏鸡和未喂食的1日龄雏鸡的肝脏细胞质中均检测到可翻译的白蛋白mRNA。可翻译的苹果酸酶mRNA仅在喂食雏鸡的肝脏中检测到。