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利用高严格度筛选程序分离编码小鼠神经生长因子γ亚基的cDNA克隆。

Isolation of a cDNA clone coding for the gamma-subunit of mouse nerve growth factor using a high-stringency selection procedure.

作者信息

Ullrich A, Gray A, Wood W I, Hayflick J, Seeburg P H

出版信息

DNA. 1984 Oct;3(5):387-92. doi: 10.1089/dna.1984.3.387.

Abstract

A new approach was taken to isolate a full-length cDNA encoding the gamma-subunit of nerve growth factor (NGF). An oligonucleotide probe was used first to identify kallikrein-related sequences in a cDNA library derived from male mouse submaxillary gland mRNA. Of 10,000 clones, 860 were identified as being kallikrein-related; these were analyzed further by hybridization to synthetic oligonucleotides using a procedure which eliminates the effects of base composition and thus allows distinction between partial and perfect sequence complementarity. Nine clones hybridized with a gamma-subunit-specific probe at high stringency, and the nucleotide sequence of the longest cDNA insert was determined. Regions of homology and variability within the kallikrein gene family are maintained in this protein and probably reflect the similar functions and different substrates for this family of proteins.

摘要

采用了一种新方法来分离编码神经生长因子(NGF)γ亚基的全长cDNA。首先使用寡核苷酸探针在源自雄性小鼠颌下腺mRNA的cDNA文库中鉴定激肽释放酶相关序列。在10,000个克隆中,860个被鉴定为与激肽释放酶相关;通过与合成寡核苷酸杂交,使用一种消除碱基组成影响从而能够区分部分和完全序列互补性的方法对这些克隆进行进一步分析。九个克隆在高严谨度下与γ亚基特异性探针杂交,并确定了最长cDNA插入片段的核苷酸序列。激肽释放酶基因家族内的同源性和变异性区域在该蛋白质中得以保留,这可能反映了该蛋白质家族的相似功能和不同底物。

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