Isolation of cDNA clones for the chicken neural cell adhesion molecule (N-CAM).
Isolation of cDNA clones for the chicken neural cell adhesion molecule (N-CAM).
复制标题
鸡神经细胞粘附分子 (N-CAM) cDNA 克隆的分离。
DOI：
10.1073/pnas.81.17.5584
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发表时间：
1984
影响因子：
11.1
通讯作者：
Cunningham,BA
中科院分区：
文献类型：
--
作者：
Murray,BA;Hemperly,JJ;Gallin,WJ;MacGregor,JS;Edelman,GM;Cunningham,BA
从9天鸡胚脑中提取神经细胞粘附分子（N-CAM）的mRNA，用抗N-CAM的抗体免疫沉淀多聚核糖体。该mRNA在兔网织红细胞裂解物系统中编程N-CAM多肽链的体外翻译。两个独立的N-CAM cDNA克隆（命名为pEC 001和pEC 020）来自富集的RNA。通过杂交选择实验验证了pEC 001对N-CAM mRNA的特异性。两种质粒均与鸡胚脑中poly（A）+ RNA中两种离散的6- 7-内切酶长RNA种类杂交，并与少量较小尺寸的多分散材料（可能是大RNA的降解产物）杂交。未检测到与胚胎肝的poly（A）+ RNA的杂交。用pEC 001进行的Southern印迹实验在用几种不同的限制性内切酶消化的鸡基因组DNA中仅检测到一个杂交片段，这表明与N-CAM mRNA的该区域内的那些序列对应的序列在鸡基因组中最多仅存在几次，并且可能仅存在一次。
Enriched mRNA coding for the neural cell adhesion molecule (N-CAM) was prepared from 9-day embryonic chicken brains by immunoprecipitation of polysomes with antibodies to N-CAM. This mRNA programmed the translation in vitro of N-CAM polypeptide chains in a rabbit reticulocyte lysate system. Two independent N-CAM cDNA clones (designated pEC001 and pEC020) were derived from the enriched RNA. The specificity of pEC001 for N-CAM mRNA was verified by hybrid selection experiments. Both plasmids hybridized to two discrete 6- to 7-kilobase-long RNA species in poly(A)+ RNA from embryonic chicken brain and to lesser amounts of polydisperse material of smaller sizes (probably degradation products of the large RNAs). No hybridization was detected to poly(A)+ RNA from embryonic liver. Southern blotting experiments with pEC001 detected only one hybridizing fragment in chicken genomic DNA digested with several different restriction enzymes, suggesting that sequences corresponding to those within this region of N-CAM mRNA are present at most only a few times, and possibly only once, in the chicken genome.