巴马小型猪主要固有免疫分子的克隆及序列分析

Cloning and sequence analysis of the main innate immune molecules in Bama miniature pig

为了开展巴马猪的固有免疫相关研究,试验采用巴马小型猪的外周血分离淋巴细胞,提取总RNA,利用特异性引物进行RT-PCR,将克隆片段与pc DNA3.1载体相连接,转化到大肠杆菌DH5α中,筛选阳性克隆并进行测序,并将测序结果提交NCBI,比较克隆序列与已知序列的同源性。结果表明:克隆的巴马猪TLR7、TLR8、TLR9、VISA、STING和IRF3序列长度分别为3 075,3 087,3 097,1 575,1 260,1 137 bp,测序结果与预期相符,各序列与猪的同源性较高。

To carry out the related research of innate immunity in Bama miniature pig, the lymphecytes were separated from the peripheral blood of Bama miniature pig, and then were used to extract the total RNA. The specific primers were used for RT - PCR, and the cloned fragments were connected with pcDNA3.1 vector, and then were transformed into E. coli DHSct. The positive clones were screened, and then the sequen- cing results were submitted to the NCBI, for comparing the homology between the cloned sequences and the known sequences. The results showed that the sequence lengths of/he TLR 7, TLR 8, TLR 9, VISA, STING and IRb3 genes from Bama miniature pig were 3075 bp, 3087 bp,3097 bp,1575 bp,1260 bp and 1137 bp, respectively. The sequencing results were in line with the expected lengths of target genes. Through sequence analysis, it was found that the sequence homology of these molecules had high homology with that of pigs.