本文選題：Ghrelin + 梅花鹿��； 參考：《內(nèi)蒙古農(nóng)業(yè)大學(xué)》2017年碩士論文

【摘要】：1999年Ghrelin作為一個(gè)內(nèi)在生長(zhǎng)激素促分泌素受體(Growth hormone secretagogue receptor,GHS-R)的配體被發(fā)現(xiàn),當(dāng) Ghrelin 與GHS-R結(jié)合后可以刺激生長(zhǎng)激素(Growth hormone,GH)的分泌。研究表明Ghrelin在生物體內(nèi)除能調(diào)節(jié)GH分泌外還有增加食欲、調(diào)節(jié)能量平衡和促進(jìn)胃酸分泌等作用。當(dāng)前國(guó)內(nèi)外對(duì)Ghrelin的研究主要集中在人、嚙齒動(dòng)物和家畜上,極少有梅花鹿方面的相關(guān)報(bào)道,只有本課題組獲得過梅花鹿Ghrelin cDNA的部分序列。為了得到梅花鹿Ghrelin基因cDNA的全長(zhǎng)序列,同時(shí)檢測(cè)Ghrelin在動(dòng)物體內(nèi)的表達(dá)與分布情況。本試驗(yàn)利用cDNA末端快速擴(kuò)增技術(shù)(RACE)、反轉(zhuǎn)錄-聚合酶鏈?zhǔn)椒磻?yīng)(RT-PCR)、實(shí)時(shí)熒光定量PCR(RT-qPCR)及免疫組織化學(xué)等技術(shù)來進(jìn)行研究。試驗(yàn)在本課題組已獲取的梅花鹿Ghrelin cDNA部分片段的基礎(chǔ)上進(jìn)行3'和5' RACE。結(jié)果成功克隆出了梅花鹿Ghrelin cDNA的3'及5'末端序列,通過與已知序列的拼接首次得到了 539 bp的梅花鹿Ghrelin cDNA全長(zhǎng)序列,其中3'非翻譯區(qū)(3' UTR)為128 bp,5' UTR為46 bp,開放閱讀框(ORF)為351 bp,該ORF編碼116個(gè)氨基酸殘基;RT-PCR和RT-qPCR結(jié)果顯示:GhrelinmRNA在梅花鹿食管、瘤胃、網(wǎng)胃、瓣胃、皺胃、十二指腸、空腸、回腸、盲腸、結(jié)腸、肝、胰、肺、脾、腎、卵巢、膀胱、心、輸尿管、子宮、胸腺、淋巴結(jié)、心內(nèi)膜、垂體前葉、腎上腺、下丘腦、舌和甲狀腺等組織內(nèi)均有表達(dá),其中在皺胃的表達(dá)量顯著高于其他器官(P0.05);免疫組織化學(xué)結(jié)果發(fā)現(xiàn):Ghrelin的免疫陽(yáng)性細(xì)胞在消化管壁內(nèi)分布較多,且主要定位于黏膜層和黏膜下層,在食管、皺胃、十二指腸和盲腸的肌層也可見免疫陽(yáng)性細(xì)胞;在實(shí)質(zhì)器官(肝、胰、肺、脾、腎、垂體前葉、甲狀腺和卵巢)內(nèi)也均有Ghrelin免疫陽(yáng)性細(xì)胞的分布,但相對(duì)較少。以上研究結(jié)果的獲得豐富了 Ghrelin的相關(guān)生物學(xué)信息,為深入研究Ghrelin在梅花鹿體內(nèi)的生理功能奠定了基礎(chǔ)。
[Abstract]:In 1999, ghrelin was found as a ligand of growth hormone secretagogue receptor GHS-R, which can stimulate the secretion of growth hormone (GH) when ghrelin binds to GHS-R. It has been shown that ghrelin can not only regulate GH secretion, but also increase appetite, regulate energy balance and promote gastric acid secretion in vivo. At present, researches on ghrelin mainly focus on human, rodent and domestic animals. There are few reports on sika deer. Only our team has obtained partial sequence of ghrelin cDNA of sika deer. In order to obtain the full-length cDNA of ghrelin gene of sika deer, the expression and distribution of ghrelin in animals were detected. In this study, rapid amplification technique of cDNA terminal, reverse transcription-polymerase chain reaction (RT-PCR), real-time quantitative PCR (RT-qPCR) and immunohistochemistry were used to study the results. The experiments were carried out on the basis of the partial cDNA fragments of ghrelin from sika deer obtained by our research group for 3 'and 5' RACEs. Results the 3 'and 5' terminal sequences of ghrelin cDNA of sika deer were cloned successfully. The full length of ghrelin cDNA of sika deer was obtained by splicing with known sequence for the first time. The ORF encodes 116 amino acid residues: RT-PCR and RT-qPCR. The results of RT-PCR and RT-qPCR showed that the expression of 10 ghrelin mRNA was detected in esophagus, rumen, reticulum, valve, rumen, duodenum, jejunum, ileum, cecum of sika deer. Colon, liver, pancreas, lung, spleen, kidney, ovary, bladder, heart, ureter, uterus, thymus, lymph node, endocardium, anterior pituitary lobe, adrenal gland, hypothalamus, tongue and thyroid were all expressed. The expression level in the abomasum was significantly higher than that in the other organs (P0.05A). The immunohistochemical results showed that the immunoreactive cells of 1% ghrelin were mainly located in the mucosal layer and submucous layer, and in the esophagus and abomasum. Immunoreactive cells were also found in the muscular layer of duodenum and cecum, and ghrelin immunoreactive cells were also found in parenchymal organs (liver, pancreas, lung, spleen, kidney, anterior pituitary lobe, thyroid gland and ovary). The above results enrich the biological information of ghrelin and lay a foundation for further study of ghrelin's physiological function in sika deer.
【學(xué)位授予單位】：內(nèi)蒙古農(nóng)業(yè)大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2017
【分類號(hào)】：S825

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本文編號(hào)：2017233