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富含亮氨酸的G蛋白偶联受体(leucine rich repeat containing G protein coupled receptor 8,LGR8)在哺乳类动物雄性精巢下降过程中起重要作用。本研究在前期完成的半滑舌鳎(Cynoglossus semilaevis)全基因组测序与转录组测序的基础上,通过RACE方法克隆得到半滑舌鳎LGR8基因(Gen Bank登录号:KP749830),该基因全长2 768 bp,开放阅读框(open reading frame,ORF)为2 205 bp,编码734个氨基酸,5’-非翻译区(untranslated regions,UTR)为272 bp,3’-UTR为296 bp。氨基酸序列中含有10个LRRs区域、1个LDL-A区域和1个7tm-1结构域。实时定量PCR(Real-time quantitative PCR,q RT-PCR)结果显示,在3龄性成熟鱼体中LGR8主要在雄鱼脑和性腺中表达,且显著高于其他组织(P<0.05);在雌鱼各组织中均有表达,但是大多数组织(除肠和心脏)中表达量无显著差异;在雄鱼性腺和脑中的表达量要显著高于雌鱼(P<0.05)。对雄鱼不同时期的性腺进行q RT-PCR检测结果显示,LGR8基因在鱼体发育80 d表达量开始上升,150~210 d表达量达到最大,且显著高于其他组织(P<0.05)。研究表明LGR8可能在半滑舌鳎性别决定和性腺发育中起一定作用,为半滑舌鳎性别决定机制的研究提供了一定的理论基础。
Leucine rich repeat containing G protein coupled receptor 8 (LGR8) plays an important role in the decline of male testis in mammals. In this study, based on the genome-wide sequencing and transcriptome sequencing of Cynoglossus semilaevis previously completed, the LGR8 gene (GenBank accession number: KP749830) was cloned by RACE. The full length of this gene is 2 768 bp, and the open reading frame The open reading frame (ORF) was 2 205 bp, encoding 734 amino acids. The open reading frame (ORF) was 272 bp in untranslated region (UTR) and 296 bp in 3’-UTR. The amino acid sequence contains 10 LRRs, 1 LDL-A and 1 7tm-1 domains. Real-time quantitative PCR (q RT-PCR) results showed that LGR8 was mainly expressed in male brain and gonads in 3-year-old mature fish, which was significantly higher than that in other tissues (P <0.05) However, most of the tissues (except the intestine and heart) showed no significant difference in the expression levels in the female gonads and in the brain (P <0.05). QRT-PCR analysis of gonads in different stages of male showed that the expression level of LGR8 began to increase at 80 days after fish development, reaching the maximum at 150 ~ 210 days, which was significantly higher than that of other tissues (P <0.05). Studies have shown that LGR8 may play a role in the sex determination and gonadal development of the soleus ostreatus, which provides a theoretical basis for the study of sex determination mechanism in the soleus halluraxis.