Cloning of terminal transferase cDNA by antibody screening.
Cloning of terminal transferase cDNA by antibody screening.
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通过抗体筛选克隆末端转移酶 cDNA。
DOI：
10.1073/pnas.81.18.5836
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发表时间：
1984
影响因子：
11.1
通讯作者：
Baltimore,D
中科院分区：
文献类型：
--
作者：
Landau,NR;StJohn,TP;Weissman,IL;Wolf,SC;Silverstone,AE;Baltimore,D
以含末端脱氧核苷酸转移酶的胸腺瘤为模板，在Lambda噬菌体载体lambda gt11中构建了一个cDNA文库。用抗末端转移酶抗体筛选斑块，鉴定出阳性克隆，其中部分阳性克隆经抗末端转移酶抗体免疫印迹电泳层析后可鉴定出β-半乳糖苷酶-cDNA融合蛋白。交叉杂交克隆的优势类别被确定为代表末端转移酶的cDNA.结果表明，一个代表性克隆与2200个核苷酸的mRNA杂交，在酶阳性的细胞中，而不与酶阴性的细胞杂交，并且cDNA所选择的翻译给出了末端转移酶的大小和抗原性的蛋白质.只有一小部分基因组DNA与可用的最长克隆杂交，这表明该序列在小鼠基因组中几乎是独一无二的。
A cDNA library was prepared from a terminal deoxynucleotidyltransferase-containing thymoma in the lambda phage vector lambda gt11. By screening plaques with anti-terminal transferase antibody, positive clones were identified of which some had beta-galactosidase-cDNA fusion proteins identifiable after electrophoretic fractionation by immunoblotting with anti-terminal transferase antibody. The predominant class of cross-hybridizing clones was determined to represent cDNA for terminal transferase by showing that one representative clone hybridized to a 2200-nucleotide mRNA in close-matched enzyme-positive but not to enzyme-negative cells and that the cDNA selected a mRNA that translated to give a protein of the size and antigenic characteristics of terminal transferase. Only a small amount of genomic DNA hybridized to the longest available clone, indicating that the sequence is virtually unique in the mouse genome.