荷斯坦奶牛BRCA2基因、STAT3基因SNPs檢測及其與乳房炎和產(chǎn)奶性狀的關(guān)聯(lián)分析
發(fā)布時(shí)間：2018-08-09 14:38
【摘要】：乳腺癌易感基因2(BRCA2)能在抑制惡性腫瘤的發(fā)生上發(fā)揮作用,并且擁有這個(gè)基因突變的家族具有較高的乳腺癌發(fā)生率。BRCA2蛋白主要參與DNA雙鏈損傷修復(fù)。BRCA2能通過多種機(jī)制對(duì)細(xì)胞周期、基因轉(zhuǎn)錄表達(dá)、DNA損傷修復(fù)等發(fā)揮不同的調(diào)節(jié)作用。牛的BRCA2基因定位于牛12號(hào)染色體上,全長約50kb,共有28個(gè)編碼外顯子。對(duì)BRCA基因的研究主要集中在人類以及小鼠上,而在奶牛上則未見相關(guān)報(bào)道,以BRCA基因作為影響奶牛產(chǎn)奶性狀及乳房炎的候選基因,探索其與產(chǎn)奶性狀及乳房炎的相關(guān)性,具有重要的研究價(jià)值。信號(hào)轉(zhuǎn)導(dǎo)及轉(zhuǎn)錄活化因子3(STAT3)是一類脫氧核糖核酸(DNA)結(jié)合蛋白。STAT3的基因在牛中則定位于第19號(hào)染色體上。STAT3蛋白廣泛表達(dá)于不同類型的細(xì)胞和組織中,既是一種胞質(zhì)信號(hào)因子,又是胞核轉(zhuǎn)錄因子,參與細(xì)胞生長、惡性轉(zhuǎn)化、凋亡等生理功能的調(diào)控。STAT3基因可被瘦素激活,在白介素-6信號(hào)通路中作為急性期反應(yīng)因子調(diào)控免疫響應(yīng)。本研究先利用DNAMAN軟件、在線軟件ExPASy及在線軟件PSORT II Prediction對(duì)奶牛BRCA2基因及STAT3基因進(jìn)行生物信息學(xué)分析。再以374份荷斯坦奶牛頸靜脈血樣為材料提取基因組DNA,利用基因組DNA擴(kuò)增目的基因序列,使用DNA測序技術(shù)檢測候選基因中的多態(tài)位點(diǎn)。采用PCR-RFLP和Forced PCR-RFLP技術(shù)對(duì)多態(tài)性位點(diǎn)進(jìn)行分型,最后與荷斯坦奶牛乳腺炎和產(chǎn)奶性狀進(jìn)行關(guān)聯(lián)分析,旨在篩查與奶牛多種產(chǎn)奶性狀及乳房炎相關(guān)的多態(tài)位點(diǎn),為中國荷斯坦奶牛的高效選育、種質(zhì)資源保存與利用提供遺傳學(xué)依據(jù)。主要研究結(jié)果及結(jié)論如下:1.BRCA2基因與STAT3基因生物信息學(xué)分析對(duì)BRCA2和STAT3進(jìn)行奶牛、水牛、綿羊、野馬和家鼠之間的核苷酸同源性分析,發(fā)現(xiàn)同源性大都在90%以上,說明基因在物種間有很高的保守性。對(duì)奶牛上述候選基因編碼的蛋白質(zhì)進(jìn)行結(jié)構(gòu)和功能預(yù)測,結(jié)果表明BRCA2和STAT3蛋白均為親水性蛋白,且均主要存在于細(xì)胞質(zhì)中。BRCA2蛋白包括了7個(gè)BRCA2重復(fù)域,而STAT3蛋白在第580-670氨基酸處有一個(gè)SH2結(jié)構(gòu)域。2.BRCA2基因與STAT3基因遺傳變異檢測本研究對(duì)BRCA2基因與STAT3基因的共18個(gè)外顯子和部分內(nèi)含子進(jìn)行分析檢測,最終檢測到4個(gè)SNPs位點(diǎn)。BRCA2基因的T48349A位點(diǎn)與C51808T位點(diǎn),STAT3基因的A43087G位點(diǎn)與T75353C位點(diǎn)。其中C51808T與T75353C均位于基因的CDS區(qū),且T75353C位點(diǎn)處于CDS區(qū)的3’非編碼區(qū),其余兩個(gè)位點(diǎn)則位于內(nèi)含子區(qū)。3.BRCA2基因與STAT3基因群體遺傳學(xué)分析對(duì)BRCA2基因與STAT3基因進(jìn)行群體遺傳學(xué)分析發(fā)現(xiàn),這四個(gè)SNP位點(diǎn)均是中度多態(tài)。BRCA2基因的T48349A位點(diǎn)與C51808T位點(diǎn)都不處于Hard-Weinberg平衡,而STAT3基因的A43087G位點(diǎn)與T75353C位點(diǎn)均處于Hard-Weinberg平衡。4.BRCA2基因與STAT3基因多態(tài)性與產(chǎn)奶性狀、乳房炎的關(guān)聯(lián)分析關(guān)聯(lián)分析表明BRCA2基因的C51808T位點(diǎn)與奶牛的日均產(chǎn)奶量和體細(xì)胞評(píng)分顯著相關(guān),CT型個(gè)體有較高的產(chǎn)奶量,TT型具備較低的體細(xì)胞數(shù);而STAT3基因的A43087G位點(diǎn)與乳蛋白率極顯著相關(guān),AG型個(gè)體的乳蛋白率相對(duì)較高。中國荷斯坦奶牛BRCA2基因和STAT3基因的生物信息學(xué)分析、多態(tài)位點(diǎn)檢測及其與產(chǎn)奶性狀和乳房炎的關(guān)聯(lián)分析,為進(jìn)一步研究BRCA2基因和STAT3基因的生物學(xué)功能提供了理論依據(jù),為了解其在乳房炎抗性中的作用機(jī)理提供一定的理論基礎(chǔ)。
[Abstract]:Breast cancer susceptibility gene 2 (BRCA2) can play a role in inhibiting the occurrence of malignant tumors, and the family with this gene mutation has a high incidence of breast cancer,.BRCA2 protein is mainly involved in DNA double chain damage repair,.BRCA2 can be regulated by a variety of mechanisms for cell cycle, gene transfer expression, and DNA damage repair. The BRCA2 gene of cattle was located on the bovine chromosome 12, with a total length of about 50kb, and there were 28 exons. The study on BRCA gene was mainly concentrated in human and mice, but no related reports were reported on the cows. The BRCA gene was used as a candidate gene for milk production and mastitis of dairy cows and was explored with milk production and mastitis. The signal transduction and transcription activator 3 (STAT3) is a class of DNA (DNA) binding protein.STAT3, which is located on chromosome nineteenth in cattle and is widely expressed in different types of cells and tissues, not only a cytoplasmic signaling factor but also a nuclear transcription factor. The regulation of.STAT3 gene with cell growth, malignant transformation, apoptosis and other physiological functions can be activated by leptin, and regulate the immune response as an acute phase reaction factor in the il--6 signaling pathway. This study first uses DNAMAN software, online software ExPASy and online software PSORT II Prediction to carry out biological letters to cow BRCA2 and STAT3 genes. The genomic DNA was extracted from the jugular vein blood samples of 374 Holstein cows. The sequence of the target gene was amplified by genomic DNA, and the polymorphic loci in the candidate genes were detected by DNA sequencing. The polymorphic loci were typed by PCR-RFLP and Forced PCR-RFLP technology. Finally, the mastitis and milk production of Holstein cows were made. Association analysis was conducted to screen polymorphic loci related to milk production traits and mastitis in dairy cows. It provides genetic basis for the efficient breeding of Chinese Holstein cows and the conservation and utilization of germplasm resources. The main results and conclusions are as follows: 1.BRCA2 and STAT3 gene bioinformatics analysis of dairy cows and water for BRCA2 and STAT3 The analysis of nucleotide homology between cattle, sheep, wild horses and domestic mice showed that the homology was more than 90%, indicating that the genes were highly conserved among the species. The structure and function of the protein encoded by the above candidate genes were predicted. The results showed that both BRCA2 and STAT3 proteins were hydrophilic proteins and were mainly found in the cytoplasm. The.BRCA2 protein contains 7 BRCA2 repeat domains, and the STAT3 protein has a SH2 domain.2.BRCA2 gene and the STAT3 gene genetic variation detection at the 580-670 amino acid. The 18 exons and the partial introns of the BRCA2 gene and STAT3 gene are analyzed and detected, and the T48349A loci of the.BRCA2 gene of the 4 SNPs loci are detected. The A43087G loci and the T75353C loci of the C51808T locus, the STAT3 gene, and the C51808T and T75353C are located in the CDS region of the gene, and the T75353C loci are in the 3 'non coding region of the CDS region, and the other two loci are in the genetic analysis of the.3.BRCA2 and STAT3 genes in the intron. It was found that both of these four SNP loci were not in Hard-Weinberg equilibrium between the T48349A loci and the C51808T loci of moderate polymorphic.BRCA2 gene, while the A43087G loci and T75353C loci of the STAT3 gene were both in Hard-Weinberg balanced.4.BRCA2 gene and STAT3 gene polymorphism and milk producing traits, and the association analysis of the association analysis of mastitis showed that the BRCA2 base was based on the association analysis of the mastitis The C51808T locus is significantly related to the daily milk production and somatic cell score of the dairy cows. The CT type has a higher milk production and the TT type has a lower body cell number, while the A43087G site of the STAT3 gene is closely related to the milk protein rate, and the milk protein rate of the AG type individuals is relatively high. The BRCA2 gene and STAT3 gene of the Chinese Holstein cows have a relatively high rate of milk protein. The analysis of Informatics, polymorphic loci and its association with milk production and mastitis provide a theoretical basis for further study of the biological functions of BRCA2 and STAT3 genes, and provide a theoretical basis for understanding the mechanism of its role in the resistance to mastitis.
【學(xué)位授予單位】：西北農(nóng)林科技大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2015
【分類號(hào)】：S858.23
本文編號(hào)：2174397
[Abstract]:Breast cancer susceptibility gene 2 (BRCA2) can play a role in inhibiting the occurrence of malignant tumors, and the family with this gene mutation has a high incidence of breast cancer,.BRCA2 protein is mainly involved in DNA double chain damage repair,.BRCA2 can be regulated by a variety of mechanisms for cell cycle, gene transfer expression, and DNA damage repair. The BRCA2 gene of cattle was located on the bovine chromosome 12, with a total length of about 50kb, and there were 28 exons. The study on BRCA gene was mainly concentrated in human and mice, but no related reports were reported on the cows. The BRCA gene was used as a candidate gene for milk production and mastitis of dairy cows and was explored with milk production and mastitis. The signal transduction and transcription activator 3 (STAT3) is a class of DNA (DNA) binding protein.STAT3, which is located on chromosome nineteenth in cattle and is widely expressed in different types of cells and tissues, not only a cytoplasmic signaling factor but also a nuclear transcription factor. The regulation of.STAT3 gene with cell growth, malignant transformation, apoptosis and other physiological functions can be activated by leptin, and regulate the immune response as an acute phase reaction factor in the il--6 signaling pathway. This study first uses DNAMAN software, online software ExPASy and online software PSORT II Prediction to carry out biological letters to cow BRCA2 and STAT3 genes. The genomic DNA was extracted from the jugular vein blood samples of 374 Holstein cows. The sequence of the target gene was amplified by genomic DNA, and the polymorphic loci in the candidate genes were detected by DNA sequencing. The polymorphic loci were typed by PCR-RFLP and Forced PCR-RFLP technology. Finally, the mastitis and milk production of Holstein cows were made. Association analysis was conducted to screen polymorphic loci related to milk production traits and mastitis in dairy cows. It provides genetic basis for the efficient breeding of Chinese Holstein cows and the conservation and utilization of germplasm resources. The main results and conclusions are as follows: 1.BRCA2 and STAT3 gene bioinformatics analysis of dairy cows and water for BRCA2 and STAT3 The analysis of nucleotide homology between cattle, sheep, wild horses and domestic mice showed that the homology was more than 90%, indicating that the genes were highly conserved among the species. The structure and function of the protein encoded by the above candidate genes were predicted. The results showed that both BRCA2 and STAT3 proteins were hydrophilic proteins and were mainly found in the cytoplasm. The.BRCA2 protein contains 7 BRCA2 repeat domains, and the STAT3 protein has a SH2 domain.2.BRCA2 gene and the STAT3 gene genetic variation detection at the 580-670 amino acid. The 18 exons and the partial introns of the BRCA2 gene and STAT3 gene are analyzed and detected, and the T48349A loci of the.BRCA2 gene of the 4 SNPs loci are detected. The A43087G loci and the T75353C loci of the C51808T locus, the STAT3 gene, and the C51808T and T75353C are located in the CDS region of the gene, and the T75353C loci are in the 3 'non coding region of the CDS region, and the other two loci are in the genetic analysis of the.3.BRCA2 and STAT3 genes in the intron. It was found that both of these four SNP loci were not in Hard-Weinberg equilibrium between the T48349A loci and the C51808T loci of moderate polymorphic.BRCA2 gene, while the A43087G loci and T75353C loci of the STAT3 gene were both in Hard-Weinberg balanced.4.BRCA2 gene and STAT3 gene polymorphism and milk producing traits, and the association analysis of the association analysis of mastitis showed that the BRCA2 base was based on the association analysis of the mastitis The C51808T locus is significantly related to the daily milk production and somatic cell score of the dairy cows. The CT type has a higher milk production and the TT type has a lower body cell number, while the A43087G site of the STAT3 gene is closely related to the milk protein rate, and the milk protein rate of the AG type individuals is relatively high. The BRCA2 gene and STAT3 gene of the Chinese Holstein cows have a relatively high rate of milk protein. The analysis of Informatics, polymorphic loci and its association with milk production and mastitis provide a theoretical basis for further study of the biological functions of BRCA2 and STAT3 genes, and provide a theoretical basis for understanding the mechanism of its role in the resistance to mastitis.
【學(xué)位授予單位】：西北農(nóng)林科技大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2015
【分類號(hào)】：S858.23
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，本文編號(hào)：2174397
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