人胎儿脑β-神经生长因子cDNA：5'和3'非翻译区的分子克隆

Human fetal brain beta-nerve growth factor cDNA: molecular cloning of 5' and 3' untranslated regions.

作者信息

Borsani G, Pizzuti A, Rugarli E I, Falini A, Scarlato G, Baralle F E, Silani V

机构信息

Istituto Sieroterapico Milanese Serafino Belfanti, University of Milan Medical School, Italy.

出版信息

Neurosci Lett. 1991 Jun 10;127(1):117-20. doi: 10.1016/0304-3940(91)90908-c.

DOI:10.1016/0304-3940(91)90908-c

PMID:1881607

Abstract

The mRNA of beta-nerve growth factor (beta-NGF) has been demonstrated to be present in the human brain, both in adult and fetal stages of development. However, its complete nucleotide sequence is unknown since a full-length cDNA has yet to be isolated. We report here the cloning and complete analysis of the human fetal brain beta-NGF transcript. cDNA synthesis was performed from total brain RNA and overlapping regions of beta-NGF cDNA were enzymatically amplified and sequenced. The central portion of the cDNA was amplified using primers designed on the known genomic DNA sequence. The 5' and 3' unknown regions were amplified by the anchored polymerase chain reaction (PCR) and by complementary DNA Ends-PCR respectively. The latter method is an original variation of Inverted PCR. The sequenced transcript is very similar to the most common form of beta-NGF mRNA present in the mouse central nervous system. In addition, both the 5' and 3' untranslated regions show a high degree of homology to the corresponding murine sequences.

摘要

β-神经生长因子（β-NGF）的信使核糖核酸（mRNA）已被证实在人类大脑中存在，无论是在发育的成年阶段还是胎儿阶段。然而，由于尚未分离出全长互补脱氧核糖核酸（cDNA），其完整的核苷酸序列尚不清楚。我们在此报告人类胎儿大脑β-NGF转录本的克隆及完整分析。从全脑核糖核酸（RNA）进行互补脱氧核糖核酸（cDNA）合成，并对β-NGF互补脱氧核糖核酸（cDNA）的重叠区域进行酶促扩增和测序。使用根据已知基因组脱氧核糖核酸（DNA）序列设计的引物扩增互补脱氧核糖核酸（cDNA）的中央部分。5'和3'未知区域分别通过锚定聚合酶链反应（PCR）和互补DNA末端PCR进行扩增。后一种方法是反向PCR的一种原始变体。测序的转录本与小鼠中枢神经系统中存在的β-NGF信使核糖核酸（mRNA）的最常见形式非常相似。此外，5'和3'非翻译区与相应的小鼠序列显示出高度同源性。