通过噬菌体展示筛选进行cDNA的表达克隆。
Expression cloning of cDNA by phage display selection.
作者信息
Light J, Maki R, Assa-Munt N
机构信息
The Burnham Institute, La Jolla, CA 92037, USA.
出版信息
Nucleic Acids Res. 1996 Nov 1;24(21):4367-8. doi: 10.1093/nar/24.21.4367.
Abstract
Expression cloning of a mouse kappa chain fragment has been achieved from a cDNA library by display of expressed proteins on filamentous phage and affinity selection for binding to anti-mouse Fab antibodies. Expressed proteins were anchored to the phage coat by a synthetic, anti-parallel leucine zipper, which had been selected from a semi-randomized zipper library for the ability to connect a test protein to phage. From a library of 4 x 10(6) transformants, two separate clones displaying different size cDNA inserts were recovered after four selection rounds. These results further demonstrate the utility of phage display for cDNA expression cloning.
摘要
通过在丝状噬菌体上展示表达的蛋白质并进行与抗小鼠Fab抗体结合的亲和选择，从小鼠cDNA文库中成功实现了小鼠κ链片段的表达克隆。表达的蛋白质通过一种合成的反平行亮氨酸拉链锚定在噬菌体外壳上，该亮氨酸拉链是从半随机拉链文库中筛选出来的，具有将测试蛋白连接到噬菌体的能力。从4×10⁶个转化体的文库中，经过四轮选择后回收了两个显示不同大小cDNA插入片段的独立克隆。这些结果进一步证明了噬菌体展示用于cDNA表达克隆的实用性。
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