Rogerson F M, LeHoux J G, Lefebre A, Mason J I
Cecil & Ida Green Center for Reproductive Biology Sciences, University of Texas Southwestern Medical Center, Dallas, USA.
J Mol Endocrinol. 1997 Oct;19(2):203-6. doi: 10.1677/jme.0.0190203.
Complementary DNAs encoding the hamster type 2 3 beta-hydroxysteroid dehydrogenase/delta 5-->4 isomerase were isolated from liver and kidney cDNA libraries. Nine clones were isolated containing identical coding and 3' untranslated sequences. However, six of the clones contained a 68-nucleotide stretch in the 5' untranslated region that was missing in the other three clones. Primers were designed to flank this region and the polymerase chain reaction (PCR) was performed on hamster liver and adrenal RNA. Two PCR products were amplified of the predicted molecular sizes and with the expected sequence. Primers were then designed to amplify sequences encompassing this region from hamster genomic DNA. Sequencing of the resultant PCR products demonstrated that the 68-nucleotide stretch missing in some transcripts corresponded exactly to the second of three exons identified. We conclude that the 5' untranslated region of this mRNA is transcribed from at least three exons, and that the sequence of the second of these exons is spliced out of some of the RNA transcripts.
从仓鼠肝脏和肾脏的cDNA文库中分离出编码仓鼠2型3β-羟基类固醇脱氢酶/δ5→4异构酶的互补DNA。分离出9个克隆,它们含有相同的编码序列和3'非翻译序列。然而,其中6个克隆在5'非翻译区含有一段68个核苷酸的片段,而另外3个克隆中则没有。设计引物使其位于该区域两侧,并对仓鼠肝脏和肾上腺RNA进行聚合酶链反应(PCR)。扩增出了两种预测分子大小且具有预期序列的PCR产物。然后设计引物从仓鼠基因组DNA中扩增包含该区域的序列。对所得PCR产物进行测序表明,某些转录本中缺失的68个核苷酸片段恰好对应于所鉴定的三个外显子中的第二个。我们得出结论,该mRNA的5'非翻译区至少由三个外显子转录而来,并且这些外显子中的第二个序列在一些RNA转录本中被剪接掉了。