Molecular cloning and mRNA expression pattern of Sox9 during sex reversal in orange-spotted grouper (Epinephelus coioides)
Molecular cloning and mRNA expression pattern of Sox9 during sex reversal in orange-spotted grouper (Epinephelus coioides)
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斜带石斑鱼性逆转过程中 Sox9 的分子克隆和 mRNA 表达模式
DOI：
10.1016/j.aquaculture.2010.06.019
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发表时间：
2010-08-15
期刊：
影响因子：
4.5
通讯作者：
Zhu, Zuo-Yan
中科院分区：
文献类型：
--
作者：
Luo, Yu-Shan;Hu, Wei;Zhu, Zuo-Yan
Sox 9基因是雄性生殖腺发育和性别决定的关键基因。为了研究其在橙斑石斑鱼雌性转雄性中的潜在作用，我们进行了以下研究。克隆了Sox 9基因，采用cDNA末端快速扩增法测定了Sox 9 mRNA的全长序列。Sox 9基因全长3277 bp，5'端非翻译区328 bp，3'端非翻译区1511 bp。该基因的开放阅读框为1437 bp，编码479个氨基酸。Sox 9基因包含3个外显子和2个内含子，两个内含子的起始和结束都符合“GT-AG”规则。RT-PCR分析表明，Sox 9 mRNA在成年橙斑石斑鱼的脑、肾、心、肝、肌肉、胃、肠、脾、睾丸和卵巢中均有表达。通过将含有17 α-甲基睾酮的药贴植入石斑鱼体内，成功地进行了人工性逆转。通过实时荧光定量PCR，我们发现Sox 9在卵巢期性腺中弱表达（处理前）。一旦性反转开始（处理后1周），Sox 9 mRNA表达水平显著升高。然而，在治疗后2周，Sox 9 mRNA表达水平显著下降至仅略高于治疗前的水平，在治疗后4周Sox 9 mRNA表达再次增加，并且在6和8周，其维持在接近治疗后1周的高水平。结果表明，Sox 9基因可能是启动和维持性逆转过程中雄性化的重要因素之一。(C)2010 Elsevier B. V.保留所有权利。
Sox9 is a key gene in male sex determination and gonad development. To study its potential function in the female-to-male sex reversal in orange-spotted grouper, we conducted the following studies. The Sox9 gene was cloned and full-length sequence of Sox9 mRNA was determined using the rapid amplification of cDNA ends method. The Sox9 mRNA consists of 3277 bp in size with a 328 bp 5' untranslated region and a 1511 bp 3' untranslated region. The 1437 bp opening reading frame encodes a 479 amino acid protein. The Sox9 gene contains 3 exons and 2 introns; the beginning and end of both introns conform to the "GT-AG" rule. RT-PCR analysis indicated that Sox9 mRNA was expressed in brain, kidney, heart, liver, muscle, stomach, intestine, spleen, testis and ovary in adult orange-spotted grouper. Artificial sex reversal was successfully performed by implanting a medicinal strip containing 17 alpha-methyltestosterone into the groupers. By real-time PCR, we found that Sox9 was weakly expressed in the ovary-stage gonads (before treatment). Once the sex reversal began (1 week after treatment), Sox9 mRNA expression level significantly increased. However, at 2 weeks after treatment, Sox9 mRNA expression level significantly decreased to a level that was only slightly higher than that before treatment Sox9 mRNA expression increased again at 4 weeks after treatment, and at 6 and 8 weeks, it was maintained at a high level close to that at 1 week after treatment. The results suggest that Sox9 may be one of the important factors initiating and maintaining the masculinization of orange-spotted grouper during sex reversal. (C) 2010 Elsevier B.V. All rights reserved.