Knöchel W, Grundmann U
Biochim Biophys Acta. 1977 May 3;476(1):47-56. doi: 10.1016/0005-2787(77)90284-2.
Nuclear steady-state RNA and polysomal RNA of chicken immature red blood cells were isolated and separated on formamide sucrose gradients. For comparison the distribution of 9 S globin mRNA was investigated by gradient centrifugation of 125I-labelled mRNA. The material was either pooled into two fractions (less than 20 S; greater than 20 S) and translated in an Ehrlich ascites cell-free system or each gradient fraction was analyzed by hybridization with [3H]-poly (U) or [3H]-labelled DNA complementary to purified 9 S globin mRNA (globin cDNA). In neither case could evidence be obtained for the existence of a high molecular weight RNA as a probable globin mRNA precursor. Further analysis was performed by electrophoresis of RNA on exponential polyacrylamide gels in formamide and subsequent hybridization with cDNA. The results are consistent with those of gradient centrifugation and demonstrate that the distribution of globin-coding sequences in nuclear steady state RNA corresponds to that of cytoplasmic 9 S globin mRNA.
分离鸡未成熟红细胞的核稳态RNA和多聚核糖体RNA,并在甲酰胺蔗糖梯度上进行分离。作为比较,通过对125I标记的mRNA进行梯度离心来研究9S珠蛋白mRNA的分布。将材料合并为两个部分(小于20S;大于20S),并在艾氏腹水细胞无细胞体系中进行翻译,或者通过与[3H] -聚(U)或与纯化的9S珠蛋白mRNA互补的[3H]标记DNA(珠蛋白cDNA)杂交来分析每个梯度部分。在这两种情况下,均未获得证据证明存在作为可能的珠蛋白mRNA前体的高分子量RNA。通过在甲酰胺中的指数聚丙烯酰胺凝胶上对RNA进行电泳并随后与cDNA杂交进行进一步分析。结果与梯度离心的结果一致,并表明核稳态RNA中珠蛋白编码序列的分布与细胞质9S珠蛋白mRNA的分布相对应。