Bajszár G, Samarina O P, Georgiev G P
Cell. 1976 Oct;9(2):323-32. doi: 10.1016/0092-8674(76)90122-7.
5' terminal nucleosides of nuclear pre-mRNA of Ehrlich ascites carcinoma cells were analyzed by a combination of different chromatographic methods and phosphatase treatment. The heavy nuclear pre-mRNA contains mainly unblocked triphosphorylated nucleosides at the 5' end, although some capped 5' ends could also be found. In this respect, it differs from cytoplasmic poly(A)+ mRNA which contains blocked 5' termini and no triphosphorylated ends. The 5' terminal nucleotides in pre-mRNA are pppGp and pppAp (in a ratio of 3:2). The determination of pppNp content in poly (A)+, poly(U)+, and poly (A)-(U)- fragments of RNA has been used as an approach to establish the topography of pre-mRNA. We also established that the technique for isolation of triphosphorylated 5' terminal fragments of RNA based on hydroxyapatite chromatography (Bajszár, Samarina, and Georgiev, 1974) is still valid in the presence of blocked oligonucleotides. The latter do not interfere with fragments containing free triphosphate groups. Using this technique, we showed that a small but significant portion of triphosphorylated 5' end fragments of 100 nucleotides in length contain oligo(U) sequences reacting with poly(A)-Sepharose.
通过多种色谱方法和磷酸酶处理相结合的方式，对艾氏腹水癌细胞核内前体mRNA的5'末端核苷进行了分析。重核前体mRNA在5'端主要含有未封闭的三磷酸化核苷，不过也能发现一些带帽的5'末端。在这方面，它与细胞质多聚腺苷酸加尾（poly(A)+）mRNA不同，后者含有封闭的5'末端，且没有三磷酸化末端。前体mRNA中的5'末端核苷酸是三磷酸鸟苷（pppGp）和三磷酸腺苷（pppAp）（比例为3:2）。测定RNA的多聚腺苷酸加尾（poly(A)+）、多聚尿苷酸加尾（poly(U)+）和无多聚腺苷酸加尾（poly(A)-）片段中的三磷酸核苷（pppNp）含量，已被用作确定前体mRNA拓扑结构的一种方法。我们还证实，基于羟基磷灰石色谱法（Bajszár、Samarin和Georgiev，1974年）分离RNA三磷酸化5'末端片段的技术，在存在封闭寡核苷酸的情况下仍然有效。后者不会干扰含有游离三磷酸基团的片段。利用该技术，我们发现一小部分但相当显著的长度为100个核苷酸的三磷酸化5'末端片段含有与多聚腺苷酸琼脂糖反应的寡聚尿苷酸（oligo(U)）序列。
