Lewis S A, Balcarek J M, Krek V, Shelanski M, Cowan N J
Proc Natl Acad Sci U S A. 1984 May;81(9):2743-6. doi: 10.1073/pnas.81.9.2743.
A clone encoding mouse glial fibrillary acidic protein (GFAP) was isolated from a cDNA library constructed so as to express the cloned sequences. The library was screened using a GFAP-specific polyclonal antiserum; a single bacterial colony expressing GFAP was identified. The complete sequence of the cDNA insert in this clone is presented, encompassing 2.5 kilobases and specifying greater than 97% of the GFAP amino acid sequence. The clone includes a long (1.4-kilobase) 3' untranslated region. Within the coding region, the data show extensive homology with other intermediate filament proteins, particularly in those regions predicted to be alpha-helical. RNA blot transfer experiments using the cloned GFAP cDNA probe revealed a single GFAP mRNA species of 2.7 kilobases in mouse brain. Southern blot analysis indicates the existence of at most two genes encoding GFAP in the mouse genome. The mouse GFAP probe cross-hybridizes weakly at high stringency with genomic DNA from diverse eukaryotic species.
从构建用于表达克隆序列的cDNA文库中分离出一个编码小鼠胶质纤维酸性蛋白（GFAP）的克隆。使用GFAP特异性多克隆抗血清筛选该文库；鉴定出一个表达GFAP的单一细菌菌落。给出了该克隆中cDNA插入片段的完整序列，其长度为2.5千碱基，确定了超过97%的GFAP氨基酸序列。该克隆包括一个长（1.4千碱基）的3'非翻译区。在编码区内，数据显示与其他中间丝蛋白有广泛的同源性，特别是在预测为α螺旋的区域。使用克隆的GFAP cDNA探针进行的RNA印迹转移实验显示，小鼠脑中存在一种2.7千碱基的单一GFAP mRNA物种。Southern印迹分析表明，小鼠基因组中编码GFAP的基因最多有两个。小鼠GFAP探针在高严谨度下与来自不同真核物种的基因组DNA弱交叉杂交。
