北极狐及乌苏里貉抑制素α亚基基因克隆及生物信息学分析

试验旨在克隆北极狐及乌苏里貉抑制素α（inhibin α，INHα）亚基基因并对其进行生物信息学分析。根据GenBank中犬科INHα预测mRNA序列（登录号：XM_545660.5）设计1对引物，用RT-PCR技术从北极狐及乌苏里貉的卵巢组织中扩增出INHα亚基基因，同时将其插入到克隆载体中，进行测序及生物信息学分析。测序结果表明，北极狐及乌苏里貉的INHα亚基基因CDS序列全长为1 107 bp，编码369个氨基酸。北极狐及乌苏里貉的INHα亚基基因与犬的同源性最高，分别为97.9%与97.6%。系统进化树分析表明，北极狐及乌苏里貉与犬亲缘关系较近，同时也说明INHα亚基基因在不同物种及进化过程中具有高度保守性。对INHα亚基蛋白的高级结构预测发现，由于半胱氨酸间形成的二硫键导致其采用"蝴蝶形"或"开放手"构型，其中α-螺旋形成分子的"手腕"结构，β-折叠形成分子的"手指"结构。本研究成功克隆了北极狐及乌苏里貉的INHα亚基基因，同时进行了系统的生物信息学分析，为今后研究抑制素在卵母细胞-颗粒细胞同步发育过程中的生物学功能奠定了基础。

Cloning and Bioinformatics Analysis of Inhibin α-subunit Gene of Arctic Fox and Wusuli Raccoon Dog

This study was aimed to clone the inhibin α(INHα) subunits gene of arctic fox and Wusuli raccoon dog and analyze its genetic structure with bioinformatics. According to the published sequence of INHα subunit gene (GenBank accession No. XM_545660.5), one pair of specific PCR primers were designed. The INHα subunit gene of arctic fox and Wusuli raccoon dog were amplified by RT-PCR. The target gene was linked into cloning vector to be sequenced and systemically analyzed by bioinformatics. The results showed that INHα subunit gene fragment of arctic fox and Wusuli raccoon dog included an 1 107 bp whole length CDS (encoding 369 amino acids). The INHα subunit gene of arctic fox and Wusuli raccoon dog shared 97.9% and 97.6% of similar nucleotide sequence with INHα subunit gene of dog, respectively. Phylogenetic analysis showed that the INHα subunit gene of arctic fox and Wusuli raccoon dog were genetically close to dog, and it was highly conserved in different species and evolutionary processes. The prediction of INHα subunit tertiary structure showed that the cysteine knot directs members of the INHα subunit to adopt a "butterfly-shaped" or "open hand" configuration, with the α-helix defining the "wrist" and β-sheets forming the "fingers" of the molecule. The INHα subunit gene of arctic fox and Wusuli raccoon dog were cloned successfully and analyzed by bioinformatics software, which would help to research the biological function of inhibin in the simultaneous development of oocyte-granulosa cells.