| 摘要: |
| 设计了黑鲷ghrelin 基因的特异性引物, 提取胃组织总RNA 并扩增出目的片段, 将PCR 产物克隆到pGEM-T Easy 载体, 经质粒PCR 扩增、酶切和测序鉴定重组质粒, 构建标准曲线等, 成功建立了ghrelin 基因荧光实时定量PCR 检测方法, 以准确分析ghrelin 基因在黑鲷摄食调节中的作用。运用建立的荧光实时定量PCR 方法检测了不同饥饿时段(2 天、7 天)黑鲷胃、肠、下丘脑组织中ghrelin mRNA 的表达变化。结果表明, 饥饿2 天(短期饥饿), 黑鲷胃、肠组织中ghrelin mRNA 表达显著上升(P<0.05), 下丘脑ghrelin mRNA 表达无显著变化(P>0.05); 饥饿7 天(长期饥饿), 黑鲷胃、肠组织ghrelin mRNA 表达比饥饿2 天显著下降, 但仍显著高于对照组, 下丘脑ghrelin mRNA 表达逐渐升高(P<0.05); 在同一饥饿时间内不同组织ghrelin mRNA 表达存在显著差异(P<0.05)。黑鲷不同组织ghrelin 基因的表达随不同饥饿时段呈现出不同的变化趋势, 推测ghrelin 基因参与了黑鲷摄食调节的生物学过程。
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| 关键词: 黑鲷, ghrelin, 实时定量PCR, 饥饿 |
| DOI：10.11693/hyhz200903010010 |
| 分类号: |
| 基金项目:国家“863 计划”项目, 2006AA10A402 号; 国家自然科学基金项目, 30700611 号; 广东省自然科学基金项目, 06027456 号;广东省高校优秀青年创新人才培育项目, A2YM08094; 惠州学院校立项目, C208.0211 号 |
附件 |
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| EFFECT OF DIFFERENT FASTING STAGES ON GHRELIN EXPRESSION IN BLACK SEABREAM ACANTHOPAGRUS SCHLEGELI |
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MA Xi-Lan1,2, ZHANG Yong1, LIU Yun1, HUANG Wei-Ren1, LIU Xiao-Chun1, ZHOU Li-Bin2
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1.State key Laboratory of Biocontrol, Institute of Aquatic Economic Animals and Guangdong Provincial Key Laboratory for Aquatic Economic Animals, Sun Yat-Sen University;2.Department of Life Science, Huizhou University
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| Abstract: |
| To analyze the roles of ghrelin in food intake of black seabream in detail, real-time quantitative PCR was adopted to examine the ghrelin gene expression. Total RNA was extracted from black seabream stomach tissue and the fragments of target gene were amplified by RT-PCR with special primers. PCR product was cloned via pGEM-T Easy vector and plasmid extracted. The recombinant plasmid was identified by PCR amplification and restriction enzyme analysis. Real time quantitative PCR was also applied to detect the ghrelin expression in stomach, intestine and hypothalamus tissues during different fasting stage. The results show that the expression of ghrelin increased in stomach and intestine tissue but not in hypothalamus tissue during a short period of fasting (2 days) (P<0.05) . However, during a long period of fasting (7 days), the expression decreased in stomach and intestine tissue, but still higher than that of the control (P<0.05), which was increased in hypothalamus tissue (P<0.05). Besides, the expression of ghrelin in different tissues at the same stage was different (P<0.05). The expression of ghrelin in different tissues did change in different fasting stages, suggesting that ghrelin is involved in the regulation of food intake in black seabream.
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| Key words: Black seabream Acanthopagrus schlegeli, Ghrelin, Real-time quantitative PCR, Fasting |
