应用原位突变技术构建免疫球蛋白K链可变区基因克隆及表达载体  

CONSTRUCTION OF A UNIVERSAL IMMUNOGLOBULIN K CHAIN VARIABLE REGION GENE CLONING AND EXPRESSION VECTOR APPLYING IN VITRO MUTAGENESIS

作　　者：张文祥[1] 程华旭 刘健[1] 

机构地区：[1]上海市肿瘤研究所免疫学和细胞生物学实验室

出　　处：《肿瘤》1995年第1期1-4,共4页Tumor

基　　金：上海市科委启明星基金

摘　　要：设计并合成了一对聚合酶链反应（ＰＣＲ）的通用引物，可以从小鼠杂交瘤的ｃＤＮＡ中扩增出免疫球蛋白（Ｉｇ）Ｋ链可变区（ＶＫ）基因。利用这一对引物和原位突变技术，在小鼠的一个ｇｅｎｏｍｉｃＶＫ基因的两端分别引入一个ＤＮＡ限制性内切酶的酶切位点，构建成一个ＩｇＶＫ基因的通用型克隆载体。并将ＶＫ基因片段同人ｋ链基因区基因片段重组，构建成人－鼠嵌合ｋ链基因表达载体。We have designed two oligonucleotide primers to amplify the cDNA of mouse immunoglobulin K chain variable domain(Vk) by the polymerase chain reaction( PCR). The primers were also used to convert the sequences of 5' and 3' coding region of a Vk genomic gene segment by sitedirected mutagenesis. The primers incorporated with restriction sites that allow the amplified cDNA of the variable domains to be force-cloned in this reconstructed universal vector. After recombination with human immunoglobulin K chain constant region gene segment,a chimeric human- mouse K chain gene can be coexpressed with a chimeric human-mouse heavy chain gene in mouse myeloma cell line such as SP_(2/0).

关 键 词：免疫球蛋白 嵌合抗体 原位突变 K链变区基因 

分 类 号：R392.11[医药卫生—免疫学] R730.3[医药卫生—基础医学]