Szeberènyi J, Goldenberg C J
Biochem Biophys Res Commun. 1984 Jul 18;122(1):466-71. doi: 10.1016/0006-291x(84)90499-6.
Precursor RNAs were synthesized in vitro from a plasmid in which the early region 2 (E2) of adenovirus 2 is fused to an efficient bacteriophage promoter (Salmonella phage 6). The RNAs were purified and utilized as substrates for in vitro splicing in the presence of nuclear extracts prepared from MOPC-315 mouse myeloma cells. We have shown previously (Goldenberg, C.J., PNAS, August, in press, 1984) that in vitro splicing in those extracts was accurate at the nucleotide level. We now show that: i) the new internucleotide bond at the splice junction generated in vitro is a 3',5'-phosphodiester bond; and ii) the phosphate that forms the splice between the exons is derived from the pre-mRNA.
前体RNA在体外由一个质粒合成，该质粒中腺病毒2的早期区域2（E2）与一个高效噬菌体启动子（沙门氏菌噬菌体6）融合。这些RNA被纯化，并用作在从MOPC - 315小鼠骨髓瘤细胞制备的核提取物存在下进行体外剪接的底物。我们之前已经表明（戈尔登伯格，C.J.，《美国国家科学院院刊》，8月，即将发表，1984年），在那些提取物中的体外剪接在核苷酸水平上是准确的。我们现在表明：i）体外产生的剪接连接处的新核苷酸键是一个3'，5'-磷酸二酯键；ii）形成外显子之间剪接的磷酸来自前体mRNA。
