牙鲆ras-2基因cDNA全长的克隆和表达分析
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上海海洋大学省部共建水产种质资源发掘与利用教育部重点实验室,上海海洋大学省部共建水产种质资源发掘与利用教育部重点实验室,上海海洋大学省部共建水产种质资源发掘与利用教育部重点实验室

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国家自然科学基金项目(31072201)


Full-length cDNA Cloning and Tissues Expression Analysis of ras-2 Gene from Paralichthys olivaceus
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The Key Laboratory of Exploration and Utilization of Aquatic Genetic Resources Ministry of Education,Shanghai Ocean University,The Key Laboratory of Exploration and Utilization of Aquatic Genetic Resources Ministry of Education,Shanghai Ocean University,The Key Laboratory of Exploration and Utilization of Aquatic Genetic Resources Ministry of Education,Shanghai Ocean University

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The National Natural Science Foundation of China (31072201)

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    摘要:

    Ras主要参与表皮生长因子家族调控细胞分裂的信号通路。本研究采用末端快速扩增法获得了牙鲆ras-2基因全长2384bp cDNA序列,该序列编码187个氨基酸,含有switchⅠ、switchⅡ、P-loop、CAAX box等Ras家族的结构特征,与川鲽ras-2氨基酸序列相似性最高(96.3%)。定量PCR检测显示ras-2基因在牙鲆成体的心脏、肝脏、脾脏、肾脏、鳃、肠、肌肉、鳍条和皮肤组织均有不同程度的表达,其中鳍条表达量最高,肌肉的表达量最低;在牙鲆变态前和变态初较高,随着变态进行,表达水平显著降低。RNA整体原位杂交结果显示ras-2基因主要在牙鲆变态阶段的鳍条、眼眶周围、颔、鳃、鼻孔、侧线等部位的表皮组织中表达,变态前在鳍条和体侧线表达信号最强,变态阶段E期的信号稍微减弱,主要分布在鳍条和鳃,变态高峰期(F期和G期)信号主要集中在鳍条、鳃和颔,H期表达信号主要集中在鳃和鳍条。ras-2基因表达式型显示,其可以通过调控细胞分裂的方式,参与牙鲆变态过程中眼睛移动、冠状幼鳍发育、侧线发育等多种器官的发育调控。

    Abstract:

    Ras is mainly involved in the epidermal growth factor family regulating the signaling pathway of cell proliferation.To investigate the relationship between ras expression and metamorphosis in Paralichthys olivaceus, the full-length cDNA of ras-2 gene with 2384bp was obtained by rapid amplification of cDNA ends (RACE). The ras-2 gene encodes 187 amino acids with typic structure such as switchⅠ, switchⅡ, P-loop and CAAX box, sharing 96.3% sequence similarity with another flatfish Platichthys flesus. RT-qPCR revealed that ras-2 gene was expressed universally in adult P. olivaceus. The highest expression of ras-2 was found in the fin, while the lowest expression in the muscle. During metamophosis of P. olivaceus, expressional level of ras-2 gene was decreasing significantly. The whole mount RNA in situ hybridization further showed the expression pattern of ras-2 during metamorphosis of P.olivaceus and revealed it expression mainly in the epidermal tissues locating in fin, jaw, orbit, nostril, lateral line and so on. Higher expression signals were observed mainly in fin, lateral line, jaw and gill during metamorphosis. Since Ras-MAPK pathway is well-known in regulating cell proliferation, basing on expressional pattern of ras-2 gene, we postulate that ras-2 might participate in eye migration and development of lateral line, dorsal fin and gill via regulating epidermic cell proliferation during metamorphosis of P.olivaceus.

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韦芬,何欣慰,鲍宝龙.牙鲆ras-2基因cDNA全长的克隆和表达分析[J].上海海洋大学学报,2013,22(5).
WEI Fen, HE Xin Wei, BAO Bao-long. Full-length cDNA Cloning and Tissues Expression Analysis of ras-2 Gene from Paralichthys olivaceus[J]. Journal of Shanghai Ocean University,2013,22(5).

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  • 收稿日期:2013-03-26
  • 最后修改日期:2013-06-03
  • 录用日期:2013-06-05
  • 在线发布日期: 2013-11-07
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