Cioe L, Curtis P
Proc Natl Acad Sci U S A. 1985 Mar;82(5):1367-71. doi: 10.1073/pnas.82.5.1367.
A cloned segment of mouse alpha-spectrin mRNA has been identified by immunological techniques. Double-stranded cDNA derived from spleens of anemic mice was introduced into a bacterial expression vector, pUC, and transformed Escherichia coli colonies were screened by using an antiserum to erythrocyte membrane ghost proteins. Of 17 positive colonies, 2 bound antibody to mouse spectrin, and these 2 colonies contained 750-base-pair inserts that cross-hybridized. Transfer of the 750-base-pair insert to an expression vector containing the PL promoter of phage lambda produced larger amounts of peptides that were bound by antibody to mouse spectrin. The spectrin-like peptides made in E. coli elicited antibody that reacted only with the alpha-spectrin subunit of erythrocyte membranes. This clone will be useful for the study of the structure and expression of the spectrin gene, particularly in understanding the role of spectrin in human inherited hemolytic anemias.
已通过免疫学技术鉴定出一段克隆的小鼠α-血影蛋白mRNA片段。将来自贫血小鼠脾脏的双链cDNA引入细菌表达载体pUC中，并用抗红细胞膜血影蛋白的抗血清筛选转化的大肠杆菌菌落。在17个阳性菌落中，有2个与小鼠血影蛋白抗体结合，这2个菌落含有可交叉杂交的750个碱基对的插入片段。将750个碱基对的插入片段转移到含有噬菌体λ的PL启动子的表达载体中，产生了大量能与小鼠血影蛋白抗体结合的肽。在大肠杆菌中产生的血影蛋白样肽引发的抗体仅与红细胞膜的α-血影蛋白亚基发生反应。该克隆将有助于研究血影蛋白基因的结构和表达，特别是有助于理解血影蛋白在人类遗传性溶血性贫血中的作用。
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