Galili G, Kawata E E, Cuellar R E, Smith L D, Larkins B A
Nucleic Acids Res. 1986 Feb 11;14(3):1511-24. doi: 10.1093/nar/14.3.1511.
A maize zein cDNA clone was used to synthesize mRNA with the SP6 in vitro transcription system. Although we obtained full-length transcripts of the cDNA sequence, these were inefficient templates for protein synthesis. Removal of the 5' oligo(G) sequence that was synthesized during the cDNA cloning procedure allowed efficient translation of the mRNAs in a wheat germ cell-free protein synthesis system or in Xenopus laevis oocytes. The alteration in translational efficiency did not result from an interaction of the 5' oligo(G) homopolymer tail with the 3' oligo(C) sequence, as transcripts with or without the oligo(C) tail were translated similarly in both protein synthesis systems. Ribosome interaction with the mRNA may be affected due either to the secondary structure of the oligo(G) sequence itself, or an unusual secondary structure between the oligo(G) sequence and another region in the mRNA. Synthetic oligonucleotides at the 5' end of cloned cDNA sequences may generally be inhibitory for translation of mRNAs transcribed in vitro.
一个玉米醇溶蛋白cDNA克隆用于通过SP6体外转录系统合成mRNA。虽然我们获得了cDNA序列的全长转录本,但这些是低效的蛋白质合成模板。去除在cDNA克隆过程中合成的5'寡聚(G)序列,使得mRNA在小麦胚无细胞蛋白质合成系统或非洲爪蟾卵母细胞中能够有效翻译。翻译效率的改变并非由5'寡聚(G)同聚物尾巴与3'寡聚(C)序列的相互作用导致,因为无论有无寡聚(C)尾巴的转录本在两个蛋白质合成系统中的翻译情况相似。核糖体与mRNA的相互作用可能受到寡聚(G)序列自身二级结构的影响,或者是寡聚(G)序列与mRNA中另一个区域之间异常二级结构的影响。克隆cDNA序列5'端的合成寡核苷酸通常可能会抑制体外转录的mRNA的翻译。