本文選題：香豬 + SNP　； 參考：《貴州大學(xué)》2016年博士論文

【摘要】：豬是一種重要的經(jīng)濟動物,不但能為人類提供充足的動物蛋白,還能作為優(yōu)良的人類疾病研究模型。繁殖性狀是豬最為重要的經(jīng)濟性狀之一,其中產(chǎn)仔數(shù)是衡量母豬繁殖能力高低的關(guān)鍵性狀。產(chǎn)仔數(shù)性狀不僅影響母豬的繁殖力,而且直接影響?zhàn)B豬業(yè)的經(jīng)濟效益,因此對產(chǎn)仔數(shù)性狀的研究成為當前研究的重點和熱點。產(chǎn)仔數(shù)性狀是數(shù)量性狀,由主效基因和多個微效基因所共同調(diào)控。隨著國內(nèi)外研究人員的努力,目前已成功定位了雌激素受體和促卵泡激素β亞基兩個與豬繁殖性狀相關(guān)的主效基因,此外,部分微效基因如骨橋蛋白,催乳素受體基因等也陸續(xù)被發(fā)現(xiàn),但是還有大量的微效基因有待發(fā)現(xiàn)。香豬是黔東南月亮山區(qū)農(nóng)民最主要的經(jīng)濟來源之一,其產(chǎn)仔數(shù)較低,平均只有6.7頭,但是變異范圍較大,產(chǎn)仔數(shù)從3~21頭都有,但是目前對于香豬的產(chǎn)仔數(shù)變異機理還不清楚。隨著人類基因組計劃和人類單倍型計劃的完成,高密度的芯片越來越多地應(yīng)用到畜牧業(yè)當中。因此本文采用Illumina Porcine SNP60K芯片,首次對貴州本地特色的香豬群體進行產(chǎn)仔數(shù)性狀的全基因組關(guān)聯(lián)分析和拷貝數(shù)變異研究,挖掘影響香豬產(chǎn)仔數(shù)性狀的重要候選基因以及相關(guān)的功能位點,掌握其產(chǎn)仔數(shù)變異的遺傳機理,在SNP水平和CNV水平為下一步的遺傳改良提供科學(xué)依據(jù)取得的主要研究結(jié)果包括:1、香豬產(chǎn)仔數(shù)性狀全基因組關(guān)聯(lián)分析本研究以98從江香豬為研究材料,通過Illumina Porcine SNP60K芯片對61565個SNP位點進行基因分型,采用混合線性模型對香豬產(chǎn)仔數(shù)性狀進行全基因組關(guān)聯(lián)研究,篩選與目標性狀顯著關(guān)聯(lián)的SNP位點,進一步尋找與香豬繁殖相關(guān)的候選基因。分型后的原始數(shù)據(jù)經(jīng)過嚴格的質(zhì)控之后,采用Plink軟件進行基于無關(guān)個體的全基因組關(guān)聯(lián)分析。分析結(jié)果經(jīng)Bonferroni校正之后,篩選到與香豬產(chǎn)仔數(shù)性狀顯著關(guān)聯(lián)的SNP位點25個,(P1.77E-06),潛在關(guān)聯(lián)SNP位點7個(1.77E-06P3.55E-05)。通過AS-PCR驗證,所有位點均與芯片分型結(jié)果一致。對顯著SNP位點上下游100 kb的基因進行檢索,共檢索到基因70個,其中13個包含關(guān)聯(lián)SNP位點,這些基因富集在16個顯著的GO條目中,影響病毒應(yīng)答、嗅覺受體、防御反應(yīng)、細胞外基質(zhì)、干擾素受體結(jié)合、細胞因子結(jié)合等生物學(xué)過程。篩選得到ZEB1、PDIA4、MARCKS、HDAC2、CDC42、FSH-β、PRSS21等基因與動物的繁殖有關(guān),可能是香豬產(chǎn)仔數(shù)性狀的潛在候選基因。2、香豬全基因組拷貝數(shù)變異檢測本研究基于Illumina Porcine SNP60K芯片數(shù)據(jù),采用PennCNV軟件從全基因組水平掃描香豬的CNVs,尋找不同產(chǎn)仔性能香豬之間的拷貝數(shù)變異作為分子標記,進一步從全基因水平挖掘候選基因的拷貝數(shù)變異對表型性狀的影響。通過一系列嚴格的質(zhì)控之后,共得到包含172個CNVRs的香豬全基因組拷貝數(shù)變異草圖,總長達到80.41Mb,大小約占豬基因組的2.98%。172個CNVRs中增加型97個;缺失型65個;缺失和增加共存型10個。有97個CNVRs與之前報道的完全或部分重疊,剩下的75個有可能是新發(fā)現(xiàn)的拷貝數(shù)變異區(qū)間。群體分析發(fā)現(xiàn)82個CNVRs是高產(chǎn)香豬特有的,而低產(chǎn)特有的只有13個。對CNVRs涉及的基因進行功能注釋,發(fā)現(xiàn)ADAMTS-1、AR、KIT、MED 12、PN-1和SOD1與繁殖有關(guān)且只在高產(chǎn)群體中存在拷貝數(shù)變異。此外,克隆了拷貝數(shù)變異候選基因ADAMTS-1,包含8個內(nèi)含子和9個外顯子區(qū)域。該基因第七外顯子5996位存在多態(tài)性,在香豬中以BB基因型為主,而西方豬種中以AA或AB型為主。高產(chǎn)和低產(chǎn)兩個香豬群體中的ADAMTS-1基因的拷貝數(shù)都是復(fù)制型為主,但存在拷貝數(shù)的差異,高產(chǎn)香豬群體中的拷貝數(shù)增加較多,多為三個或三個以上拷貝,而低產(chǎn)群體中多為雙拷貝,高產(chǎn)和低產(chǎn)群體中缺失型的個體分別有3頭和16頭。結(jié)果表明低產(chǎn)群體中的拷貝數(shù)低于高產(chǎn)群體。本文通過以上研究發(fā)現(xiàn)了與香豬產(chǎn)仔數(shù)性狀顯著關(guān)聯(lián)的SNP位點和候選基因,為后續(xù)候選基因的功能驗證,分子育種,揭示香豬繁殖力調(diào)控機理奠定基礎(chǔ)。首次繪制得到香豬全基因組拷貝數(shù)變異草圖,為進一步研究香豬表型差異與拷貝數(shù)變異之間的關(guān)系提供理論依據(jù)。而ADAMTS-1基因的拷貝數(shù)變異以及第七外顯子5996位的多態(tài)性可能是影響香豬繁殖力的重要因素之一。
[Abstract]:Pig is an important economic animal, which not only provides sufficient animal protein for human beings, but also can be used as a good model of human disease research. Reproductive traits are one of the most important economic traits of pigs. The number of litter is the key trait to measure the breeding ability of sows. The study of the number of litter size has become the focus and hot spot in the present study. The number of litter size traits is a quantitative trait, which is regulated by the main gene and the multiple micro effect genes. With the efforts of researchers at home and abroad, two of the estrogen receptor and the follicle stimulating hormone beta subunit have been successfully located. In addition, some micro effective genes, such as osteopontin and prolactin receptor genes, have been discovered in succession, but there are still a large number of micro effective genes to be found. The fragrant pig is one of the most important economic sources of farmers in the Qiandongnan moon mountain area, with a lower litter size and an average of only 6.7, but the range of variation is larger. The number of litter is from 3~21, but the variation mechanism of the litter size is unclear. With the completion of the human genome project and the human haplotype program, the high density chip is more and more applied to the animal husbandry. Therefore, this paper uses the Illumina Porcine SNP60K core for the first time to the local characteristics of the Guizhou pig population. Complete genome association analysis and copy number variation study of litter size traits, excavating important candidate genes and related functional sites affecting litter size traits of fragrant pigs, mastering the genetic mechanism of litter size variation, and providing scientific basis for the next step of genetic improvement at SNP level and CNV level: 1, The whole genome association analysis of the litter size traits of Xiang pig 98 from Jiangxiang pig as the research material, 61565 SNP loci were genotyping by Illumina Porcine SNP60K chip. A mixed linear model was used to study the whole genome association of the litter size traits of the fragrant pigs, and to screen the SNP site which was significantly associated with the target characters. After finding the candidate genes related to the reproduction of the pig, the original data after the typing were strictly controlled by the Plink software to analyze the whole genome association based on the unrelated individuals. After the Bonferroni correction, 25 SNP loci, which were significantly associated with the litter size traits of the pigs, were selected, (P1.77E-06), and 7 potential associated SNP loci. (1.77E-06P3.55E-05). Through AS-PCR verification, all loci were consistent with the results of the microchip typing. A total of 70 genes were retrieved for 100 kb in the upper and lower reaches of the significant SNP loci, of which 13 contained associated SNP sites, and these genes were enriched in 16 significant GO entries, affecting the viral response, the olfactory receptor, defense response, and the extracellular matrix. ZEB1, PDIA4, MARCKS, HDAC2, CDC42, FSH- beta, PRSS21 and other genes are related to the reproduction of animals. It may be a potential candidate gene for the litter size of the pig. The whole genome copy number variation detection of Xiang pig is based on Illumina Porcine SNP60K chip data. PennCNV software was used to scan the whole genome level of the CNVs of the fragrant pigs, looking for the copy number variation between the different piglets of different litter size as molecular markers, and further mining the influence of the copy number variation on the candidate genes from the whole gene level to the phenotypic traits. After a series of strict quality control, a total of 172 CNVRs pig whole genes were obtained. The total length of copy number variant sketches reached 80.41Mb, the size of which accounted for 97 of the 2.98%.172 CNVRs in the pig genome, 65 missing, 10 missing and increased coexistence. 97 CNVRs were fully or partially overlapped with previous reports, and the remaining 75 may be the newly discovered copy number variation interval. Group analysis found that 82 CNVRs was high. There are only 13 endemic pigs and only 13 low yield traits. Functional annotation of genes involved in CNVRs found that ADAMTS-1, AR, KIT, MED 12, PN-1 and SOD1 are related to reproduction and have copy number variation only in high yield populations. In addition, the copy number variant candidate gene ADAMTS-1, including 8 introns and 9 exons, has been cloned. Seven exon 5996 was polymorphic, and BB genotypes were dominant in fragrant pigs, while AA or AB type was dominant in Western pigs. The number of copies of ADAMTS-1 genes in two high and low yield pigs were replicative, but there was a difference in copy number. The number of copies in high yield pigs increased more, more than three or more than three copies. There are more than two copies in the low yield population and 3 and 16 in the high and low yield individuals. The results show that the number of copies in the low yield population is lower than that of the high yield population. In this paper, the SNP loci and candidate genes associated with the litter size traits of the pigs were found, and the functional verification of the subsequent candidate genes was found. Seed breeding is the basis for revealing the regulation mechanism of the reproductive power of the pig. The whole genome copy number variant sketch of the Xiang pig is first drawn to provide a theoretical basis for further study of the relationship between the phenotypic and copy number variations of the Xiang pig. The variation of the copy number of the ADAMTS-1 gene and the polymorphism of the seventh exon 5996 may affect the fragrant pigs. One of the important factors of fecundity.
【學(xué)位授予單位】：貴州大學(xué)
【學(xué)位級別】：博士
【學(xué)位授予年份】：2016
【分類號】：S828

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