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圆斑星鲽sox9基因的克隆与表达
张乐乐1,2, 边力2, 常青2, 侯吉伦3, 陈四清2, 赵庆1,2, 刘琨1,2, 葛建龙2, 刘长琳2
1.水产科学国家级实验教学示范中心 上海海洋大学 上海 201306;2.中国水产科学研究院黄海水产研究所 青岛 266071;3.中国水产科学研究院北戴河中心实验站 秦皇岛 066100
摘要:
本研究筛选到圆斑星鲽(Verasper variegatus)的性别相关基因sox9,并通过RACE技术获得了全长序列,基因全长为3287 bp,包括1431 bp的ORF,编码477个氨基酸,368 bp的5¢ UTR和1488 bp的3¢ UTR。在3¢ UTR中有多聚腺苷酸尾和加尾信号AATAAA。通过荧光定量PCR测定了sox9基因在圆斑星鲽成鱼不同组织中的表达水平,发现sox9基因在圆斑星鲽的脑、眼、鳃、心、肝、胆、肠、精巢、卵巢、肾和肌肉等各个组织中都有不同程度的表达。在鳃、脑和精巢组织中检测到较高水平的sox9转录,其中精巢中的转录水平显著高于其他组织,sox9基因在性腺中的表达显示出性别两相性差异。其在精巢中的表达水平要显著高于卵巢,说明sox9基因与雄性性腺发育相关。通过测定sox9基因在圆斑星鲽幼鱼不同发育时期(20、30、40、50、60、70和80日龄)的表达水平,发现其在20~50日龄表达量逐渐下降,在60日龄时表达量上升,推测表达量上升可能与幼鱼性腺分化相关。
关键词:  圆斑星鲽  sox9  性别基因  mRNA表达
DOI:
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基金项目:
Cloning and Expression of the sox9 Gene in Spotted
ZHANG Lele1,2, BIAN Li2, CHANG Qing2, HOU Jilun3, CHEN Siqing2, ZHAO Qing1,2, LIU Kun1,2, GE Jianlong2, LIU Changlin2
1.National Demonstration Center for Experimental Fisheries Science Education, Shanghai Ocean University, Shanghai 201306;2.Yellow Sea Fisheries Research Institute, Chinese Academy of Fishery Sciences, Qingdao 266071;3.Beidaihe Central Experiment Station, Chinese Academy of Fishery Sciences, Qinhuangdao 066100
Abstract:
Significant differences exist in the growth rate and body size of male and female spotted halibut (Verasper variegatus), with the females being significantly larger than the males. All-female cultivation can improve breeding efficiency. Studies investigating the mechanism of sex determination and features of sex differentiation may provide valuable information for all-female cultivation. In this study, we successfully identified the sox9 gene of spotted halibut. The total length is 3287 bp, including a 1431 bp ORF, encoding 477 amino acids; the 5¢ UTR is 368 bp and the 3¢ UTR is 1488 bp. Polyadenylic acid tails and AATAAA caudal signals were identified in the 3¢ UTR. The expression of the sox9 gene differed in the brain, eye, gills, heart, liver, gallbladder, intestine, testis, ovary, kidney, and muscle in spotted halibut. Transcript levels were high in the gill, brain, and testis, and were significantly higher in the testis than in the other tissues. A two-phase gender difference was observed in the gonads; sox9 expression was significantly higher in the testis than in the ovary, indicating that sox9 might be associated with male gonad development. The dynamic expression patterns of the sox9 gene in spotted halibut larvae gradually declined from 20 to 50 d after hatching, with a sudden rise observed after 60 d. This increased expression might be associated with differentiation of larval gonad.
Key words:  Verasper variegatus  sox9  Sex genes  mRNA expression