牛胚系基因中两种JH和Cμ抗体基因的克隆与分析

Cloning and Characterization of Two JH and CμIg Genes in Bovine Germline Gene

根据NCBIGenBankTM中登录的2个牛抗体重链可变区J基因(JH)(序列号为AY158087,AY149283)的序列不同之处设计PCR引物,能从同一个体的Holstein牛基因组DNA中扩增得到与以上2个基因分别相同的序列,证明这2个JH基因的差异不是由于牛个体或品种不同引起的.提取牛脾脏总RNA,RT-PCR扩增IgMcDNA,测序结果显示:序列号为AY149283的JH基因中第六外显子JH6是一个功能基因,它可以编码牛IgM的部分CDR3区和完整的FR4区,从2种IgMcDNA克隆的测序结果可以看出,其恒定区序列分别与序列号为U63637和AY230207的2种抗体重链恒定区μ基因(Cμ)cDNA序列相同.PCR扩增JH-Cμ序列,测序结果表明:序列号为AY158087的JH基因是同以上2种Cμ基因分别串连在一起的(序列号分别为AY230207和U63637).以上结果证明:序列号为U63637和AY230207的2个Cμ基因分别与AY149283的JH基因串联在一起,都能够参与牛IgM的产生,它们与AY158087的JH基因共同存在于同一个体Holstein牛胚系基因中.

Using methods based upon the PCR, two DNA sequences which are identical to two JH genes （accession No.AYI58087, AY149283） from one Holstein bovine genomic DNA have been cloned, the results demonstrate that all the two JH genes（accession No.AYI58087, AY149283） are present in the same Holstein bovine genomic DNA. Two kinds of IgM cDNA was cloned from bovine spleen total RNA. Sequencing the cDNA clones show that the last exon of JH gene （accession No.AY149283） JH6 is a functional gene encoding part of CDR3 and whole FR4 region of bovine IgM. The sequences of two IgM cDNA clones indicate each constant region is identical to one of the two Cμ. genes（accession No.AY230207, U63637）, there are greater diversities in the IgM CH1 domain than the other CH domains. PCR products of JH-Cμ. gene suggested that each Cμ. gene （accession No.AY230207, U63637） is in series with the same JH gene （accession No. AY149283） in the bovine germline genes, they are all functional genes in the process of bovine IgM generation.