Lindberg R A, Thompson D P, Hunter T
Molecular Biology and Virology Laboratory, Salk Institute, San Diego, California 92138.
Oncogene. 1988 Dec;3(6):629-33.
A phage lambda gt11 human fibroblast cDNA expression library was screened with antibodies against phosphotyrosine to determine the feasibility of this approach as a method for the identification of clones that code for protein-tyrosine kinases. Several antibody positive clones were isolated. One clone also scored positive with degenerate oligonucleotides designed to identify sequences coding for protein-tyrosine kinases. This cDNA clone was partially sequenced and proved to be identical to part of lyn, a recently reported putative protein-tyrosine kinase. A portion of the cDNA was cloned into an inducible plasmid expression vector. Phosphorylation of many bacterial proteins on tyrosine was observed upon addition of inducing agent. The results demonstrate that this method of screening can identify cDNAs that encode active protein-tyrosine kinases and could prove useful for the identification of novel ones.
用抗磷酸酪氨酸抗体筛选λ噬菌体gt11人成纤维细胞cDNA表达文库，以确定这种方法作为鉴定编码蛋白酪氨酸激酶的克隆的方法的可行性。分离出几个抗体阳性克隆。一个克隆用设计用于鉴定编码蛋白酪氨酸激酶的序列的简并寡核苷酸也呈阳性。对该cDNA克隆进行了部分测序，结果证明它与lyn的一部分相同，lyn是最近报道的一种假定的蛋白酪氨酸激酶。将部分cDNA克隆到诱导型质粒表达载体中。加入诱导剂后，观察到许多细菌蛋白在酪氨酸上发生磷酸化。结果表明，这种筛选方法可以鉴定编码活性蛋白酪氨酸激酶的cDNA，并且可能对鉴定新的蛋白酪氨酸激酶有用。
