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豚鹿MHC-DOA基因克隆及其外显子2遗传多样性研究
Cloning of Axis porcinus MHC-DOA Gene and the Genetic Diversity of Its Exon 2 Region
邵欢欢1,2#, 邓家波3#, 吴林峰1, 牛李丽3, 余建秋3, 谭雪梅1*
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DOI:10.11984/j.issn.1000-7083.20190091
作者单位:1. 四川大学生命科学学院, 生物资源与生态环境教育部重点实验室, 四川省分子生物学及生物技术重点实验室, 成都 610065;
2. 四川师范大学生命科学学院, 成都 610101;
3. 成都动物园, 成都 610081
中文关键字:豚鹿;MHC-DOA;外显子2;基因多态性
英文关键字:Axis porcinus; MHC-DOA; exon 2; gene polymorphism
中文摘要:主要组织相容性复合物(MHC)在脊椎动物的免疫系统中起着重要的作用,MHC-DOA基因是MHCⅡ类的非经典基因,其外显子2多态性丰富。豚鹿Axis porcinus是我国极度濒危的动物。本研究以成都动物园圈养的38头豚鹿为研究对象,提取豚鹿血液总RNA,反转录合成cDNA为模板,利用PCR方法克隆到豚鹿MHC-DOA序列;同时利用PCR方法扩增38头豚鹿的MHC-DOA基因外显子2基因并测序,再进行多态性分析。结果显示:豚鹿MHC-DOA基因开放阅读框长753 bp,编码250个氨基酸残基;蛋白质同源性分析表明,豚鹿MHC-DOA基因与东欧马鹿Cervus elaphus hippelaphus的同源性最高(98.4%);38头豚鹿的MHC-DOA外显子2共有10种单倍型,整体遗传多样性水平中等;MHC-DOA外显子2部分序列中发生了核苷酸的缺失和插入,进而引起DOA蛋白序列发生改变。豚鹿MHC-DOA多态性的研究对豚鹿种群遗传结构调查、遗传资源的保护具有重要意义。
英文摘要:The genes encoding the major histocompatibility complex (MHC) are important in the immune system of vertebrates. MHC-DOA gene is a non-classical gene of MHC class Ⅱ, of which exon 2 has rich polymorphisms. Hog deer (Axis porcinus) is an extremely endangered species in China. In this study, total RNA was extracted from the blood of A. porcinus and then the MHC-DOA gene was cloned by PCR from cDNA template. Meanwhile, the exon 2 of MHC-DOA genes in 38 A. porcinus from Chengdu Zoo were obtained by PCR and DNA-sequencing, followed by genetic polymorphism analysis. The results showed that the MHC-DOA gene of A. porcinus contained a 753 bp open reading frame, encoding a protein with a length of 250 amino acid residues. Sequence analysis predicted that DOA protein had high amino acids similarity (98.4%) with Cervus elaphus hippelaphus. A total of 10 haplotypes of DOA exon 2 were detected from the 38 individuals, indicating a medium level of genetic diversity in A. porcinus. Nucleotides insert and deletion were observed in MHC-DOA exon 2 gene, and these resulted in the change of MHC-DOA amino acid sequence. This study has significant importance in understanding the population genetic structure and the protection of A. porcinus in the future.
2020,39(1): 81-87 收稿日期:2019-03-14
分类号:Q959.842;Q987.1
基金项目:成都大熊猫繁育研究基金项目(CPF-2010-02)
作者简介:邵欢欢(1984-),男,博士,讲师,从事分子遗传学研究,E-mail:wsshforget@163.com;邓家波(1974-),男,硕士,高级兽医师,从事野生动物的研究,E-mail:565211989@qq.com
*通信作者:谭雪梅,讲师,主要从事动物分子遗传学研究,E-mail:txmyyf@scu.edu.cn
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