Esnault R, Trapy G, van Huystee R B
Eur J Biochem. 1975 Dec 1;60(1):83-9. doi: 10.1111/j.1432-1033.1975.tb20978.x.
After incubating root apices from two-day-old bean seedlings with [3H] adenine the RNA was extracted from whole cells or polysomes, and the poly (A) sequences were isolated by nuclease digestion followed by poly(U)-Sepharose chromatography. The alterations of the RNA molecules due to the various treatments were monitored by sucrose density gradients. It was found that sequential extraction first at pH 7.6 then at pH 9.0 did not result in a separation between RNA poor in poly(A) sequences and poly(A)-rich RNA. Furthermore chromatography analysis of hydrolysates from nuclease-resistant RNA extracted either at pH 7.6 or pH 9.0 revealed that AMP constituted nearly 95% of the bases and that the poly(A) sequences, about 200 bases, were located at the 3' terminus of the polyadenylated RNA. No size difference was found for the poly(A) segment between the pH-7.6-extracted RNA and that extracted at pH 9.0.
用[3H]腺嘌呤培养两天大的菜豆幼苗的根尖后,从全细胞或多核糖体中提取RNA,通过核酸酶消化,随后用聚(U)-琼脂糖凝胶柱色谱法分离聚(A)序列。用蔗糖密度梯度监测各种处理导致的RNA分子变化。结果发现,先在pH 7.6然后在pH 9.0进行连续提取,并不会使富含聚(A)序列的RNA与缺乏聚(A)序列的RNA分离。此外,对在pH 7.6或pH 9.0提取的核酸酶抗性RNA水解产物的色谱分析表明,AMP占碱基的近95%,并且聚(A)序列(约200个碱基)位于聚腺苷酸化RNA的3'末端。在pH 7.6提取的RNA和在pH 9.0提取的RNA之间,未发现聚(A)片段存在大小差异。