关键词: 外周血, 免疫球蛋白, 轻链, 可变区, 反转录聚合酶链式反应, 兔

Abstract: Objective To amplify immunoglobulin Kappa light chain variable genes rapidly from rabbit peripheral blood. Methods PCR primers were designed according to the cDNA sequence of rabbit germline IGKV, IGKJ and IGKC genes, which were obtained from IMGT/GENE-DB. With the total RNA extracted from rabbit peripheral blood as template, an one-step RT-PCR reaction was done, and the products were cloned to T vector, sequenced, compared with rabbit germline Kappa light chain variable genes in IMGT/V-QUEST. By counting the kinds of Vκ-Jκ-Cκ assembly, the specificity and compatibility of the primers were evaluated. Results Of 4 pairs of primers designed, RK.C1［1-2-7］, RK.C1［3-6-8］and RK.C2［1-2-3-4］, but not RK.C1［4-5-9］were succeeded in amplifying the rabbit total RNA. Fifty-one clones from their products all belonged to rabbit immunoglobulin Kappa light chain variable and constant genes and different each other. Among them, 33 clones’constant regions were encoded by IGKC1*1, the rest, 18 clones, were encoded by IGKC2*03, and up to 22 of 68 IGKV and only 1 of 8 IGKJ functional germline genes appeared. Totally, 17 kinds of Vκ-Jκ-IGKC1 and 11 kinds of Vκ-Jκ-IGKC2 assembly were found in these 51 clones. IGKV1S10*01 was found in 12 clones, including 7 Vκ-Jκ-IGKC1 and 5 Vκ-Jκ-IGKC2 clones, which was very higher than other Vκ genes； There were existed some differences in sequence, although the 12 clones were from the same Vκ germline ge

Key words: Peripheral blood, Immunoglobulin, Light chain, Variable region, RT-PCR, Rabbit