本文關(guān)鍵詞： 黃顙魚 GH基因 單核苷酸多態(tài)性 微衛(wèi)星 生長性狀　出處：《華南農(nóng)業(yè)大學(xué)》2016年碩士論文　論文類型：學(xué)位論文

【摘要】：本研究對黃顙魚生長激素基因全長進(jìn)行克隆測序,分析GH在組織中的差異性表達(dá),對GH基因的單核苷酸多態(tài)性(SNP)和微衛(wèi)星(SSR)多態(tài)性與主要的生長性狀進(jìn)行關(guān)聯(lián)分析,并獲得調(diào)控經(jīng)濟(jì)性狀的優(yōu)勢基因位點(diǎn)。(1)GH基因的分子特征根據(jù)親緣關(guān)系較近物種的GH基因的保守區(qū)域設(shè)計(jì)引物,通過克隆測序拼接后獲得DNA全長序列,共3117bp,包括5′UTR 165bp,3′UTR 485bp,由5個(gè)外顯子和4個(gè)內(nèi)含子組成,外顯子大小分別為10bp、140bp、117bp、132bp、204bp,內(nèi)含子大小分別為302bp、85bp、642bp、835bp。(2)GH基因的差異性表達(dá)通過半熒光定量和實(shí)時(shí)熒光定量兩種方法研究黃顙魚GH基因mRNA的差異性表達(dá)。結(jié)果顯示:GH基因只在垂體中特異表達(dá),而在肌肉、脾臟、脂肪、腎臟、胃、盲腸、心臟、頭腎、精巢、腸、鰓、肝臟、魚皮和腦這14個(gè)組織中均未檢測到表達(dá)。(3)SNP與生長性狀的關(guān)聯(lián)性以GH基因作為影響黃顙魚主要生長性狀的候選基因,樣本數(shù)為160,通過PCR產(chǎn)物直接測序法篩選SNP位點(diǎn),篩選區(qū)域?yàn)槠瓮怙@子4到片段外顯子5和3′UTR區(qū)域,共檢測到5個(gè)SNP位點(diǎn),內(nèi)含子4區(qū)域檢測到4個(gè),3′UTR發(fā)現(xiàn)1個(gè)。并對SNP位點(diǎn)基因型和主要生長性狀(體重、全長、體長、體高、體厚、頭長、尾柄長、尾柄高)進(jìn)行關(guān)聯(lián)分析,結(jié)果顯示:存在于內(nèi)含子4的位點(diǎn)1674GA和位點(diǎn)2154TG與生長性狀無顯著關(guān)聯(lián);位點(diǎn)2100AG與性狀體厚有極顯著關(guān)聯(lián)(P0.01),GG為優(yōu)勢基因型;位點(diǎn)2285TC與尾柄長有顯著關(guān)聯(lián)(P0.05),CC為優(yōu)勢基因型;存在于3′UTR的位點(diǎn)2822AG與全長、尾柄高有顯著關(guān)聯(lián)(P0.05),與體長、尾柄長有極顯著關(guān)聯(lián)(P0.01),GG為優(yōu)勢基因型。(4)SSR與生長性狀的關(guān)聯(lián)性通過SSRHunter軟件在黃顙魚GH基因上查找到4個(gè)微衛(wèi)星位點(diǎn),將其在96-毛細(xì)管ABI3730xl上進(jìn)行檢測,根據(jù)檢測結(jié)果判定基因型,并與生長性狀進(jìn)行關(guān)聯(lián)分析。結(jié)果表明:位點(diǎn)AT與生長性狀無顯著關(guān)聯(lián);位點(diǎn)GA與體重、體長、體高、體厚有顯著關(guān)聯(lián)(P0.05),與全長、尾柄高有極顯著關(guān)聯(lián)(P0.01),CD為優(yōu)勢基因型;位點(diǎn)TCTT與頭長、尾柄高有顯著關(guān)聯(lián)(P0.05),BD為優(yōu)勢基因型;位點(diǎn)AC與體厚、頭長有顯著關(guān)聯(lián)(P0.05),DD為優(yōu)勢基因型。
[Abstract]:In this study, the full-length growth hormone gene of Pelteobagrus fulvidraco was cloned and sequenced, and the differential expression of GH in tissues was analyzed. Single nucleotide polymorphisms (SNPs) and microsatellite SSRs (SSRs) of GH gene were associated with major growth traits. The molecular characteristics of the GH gene were designed according to the conserved region of the GH gene of the nearest species. The full-length sequence of DNA was obtained by cloning and sequencing, which consisted of 5 exons and 4 introns. The size of exon is 10bp, 140bp, 117bp, 132bp, 204bp, and the intron is 302bp, 85bp, 642bp, respectively. 835bp. / 2). The differential expression of GH gene in Pelteobagrus fulvidraco was studied by semi-fluorescent quantitative method and real-time quantitative method. The results showed that the specific expression of the GH gene was found only in pituitary gland of Pelteobagrus fulvidraco. But in muscle, spleen, fat, kidney, stomach, cecum, heart, head kidney, testis, intestine, gills, liver. There was no correlation between the expression of SNP and growth traits in fish skin and brain. The GH gene was used as the candidate gene to affect the main growth traits of Pelteobagrus fulvidraco. The sample number was 160. The SNP loci were screened by direct sequencing of PCR products. Five SNP loci were detected in the regions from exon 4 to exon 5 and exon 3. SNP locus genotypes and main growth traits (body weight, full length, body length, body height, body thickness, head length, tail stalk length) were detected in intron 4 region. The results showed that there was no significant correlation between locus 1674GA and locus 2154TG in intron 4. Locus 2100AG was significantly associated with the body thickness of the trait P0.01GG was the dominant genotype. Locus 2285TC was significantly associated with the length of caudal stalks. P0.05C was the dominant genotype. There was a significant correlation between the height of the tail stipe and the body length, the length of the tail stalk and the length of the tail stipe (P0.01). Four microsatellite loci were found on the GH gene of Pelteobagrus fulvidraco by SSRHunter software. It was detected on 96- capillary ABI3730xl, and genotypes were identified according to the detection results. The results showed that there was no significant correlation between AT locus and growth traits. There was a significant correlation between GA and body weight, body length, body height, body thickness, and total length. There was a significant correlation between TCTT locus and head length and height of caudal stalks (P0.05BD) as dominant genotypes. There was a significant correlation between AC and body thickness and head length.
【學(xué)位授予單位】：華南農(nóng)業(yè)大學(xué)
【學(xué)位級別】：碩士
【學(xué)位授予年份】：2016
【分類號】：S917.4

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