綿羊ANGPTL4基因克
發(fā)布時(shí)間：2018-09-08 12:06
【摘要】：血管生成素樣蛋白4(ANGPTL4)兼有促進(jìn)脂肪沉積和脂肪酸氧化的作用。一方面,它通過抑制脂蛋白脂肪酶活性而抑制脂肪水解,導(dǎo)致脂代謝相關(guān)疾病的發(fā)生。另一方面,通過加強(qiáng)兒茶酚胺或糖皮質(zhì)激素的活性而促進(jìn)脂肪分解。目前,對(duì)于該蛋白及其編碼基因的研究已有報(bào)道,但在綿羊中未見報(bào)道。本研究在DNA.RNA和蛋白三個(gè)水平上對(duì)ANGPTL4基因的結(jié)構(gòu)和功能,mRNA和蛋白的表達(dá)以及ANGPTL4基因5’非編碼區(qū)的多態(tài)性進(jìn)行了研究,并與相關(guān)性狀進(jìn)行了關(guān)聯(lián)分析。利用RT-PCR技術(shù)克隆了綿羊ANGPTL4基因的編碼區(qū)序列(coding sequence, CDS),用生物信息學(xué)方法分析了CDS區(qū)及其編碼的氨基酸序列。以具有顯著脂尾型差異的脂尾型綿羊：廣靈大尾羊和小尾寒羊,共96只為研究對(duì)象,用實(shí)時(shí)熒光定量PCR(Quantitative real-time PCR)技術(shù),檢測(cè)了ANGPTL4 mRNA在2、4、6、8、10、12,共6個(gè)年齡組的7種脂肪組織(大網(wǎng)膜、小網(wǎng)膜、皮下、腹膜后、’腎周、腸系膜以及尾部)中的相對(duì)表達(dá)量,并與兩品種的尾性狀和屠宰性狀進(jìn)行了關(guān)聯(lián)研究。用Western blotting技術(shù)檢測(cè)了ANGPTL4蛋白在兩品種母羊的4種脂肪組織(皮下、腹膜后、腎周以及尾部)中的相對(duì)表達(dá)量。用PCR-SSCP技術(shù)檢測(cè)了96只個(gè)體的ANGPTL4基因5’非編碼區(qū)的單核苷酸多態(tài)性(SNP),并與相關(guān)性狀進(jìn)行了關(guān)聯(lián)研究。主要結(jié)果如下：(1)綿羊ANGPTL4基因CDS區(qū)長1233 bp,編碼410個(gè)氨基酸。預(yù)測(cè)的ANGPTL4蛋白的等電點(diǎn)為9.02,屬于堿性蛋白,相對(duì)分子量45.5 kDa。該蛋白是一種不穩(wěn)定的親水蛋白。作為一個(gè)分泌蛋白,它含有一個(gè)長為24 AA的信號(hào)肽,3次跨膜。氨基酸序列中有一個(gè)N-糖基化位點(diǎn)和一個(gè)O-糖基化位點(diǎn),18個(gè)磷酸化位點(diǎn)。進(jìn)化分析表明,綿羊與牛的親緣關(guān)系最近。(2)ANGPTL4 mRNA在廣靈大尾羊和小尾寒羊兩品種中存在時(shí)空表達(dá)差異。品種、月齡、組織對(duì)ANGPTL4 mRNA表達(dá)影響極顯著(P0.001)。在廣靈大尾羊中的表達(dá)量顯著高于小尾寒羊。10月齡時(shí)的表達(dá)量顯著高于其它月齡,淺層脂肪組織中的表達(dá)量高于在深層脂肪組織。性別影響不顯著但是公羊表達(dá)量稍低于母羊。品種與月齡(P0.001)、品種與組織(P0.01)和性別與組織(P0.01)間的互作也影響ANGPTL4mRNA的表達(dá)。mRNA表達(dá)量與屠宰和尾型性狀之間的相關(guān)性因品種差異而不同。廣靈大尾羊各組織中的表達(dá)量均與性狀呈正相關(guān),其中,皮下、腹膜后、大網(wǎng)膜和小網(wǎng)膜中的表達(dá)量與各性狀的相關(guān)顯著。小尾寒羊大網(wǎng)膜和腹膜后脂肪中的表達(dá)量與大多數(shù)性狀呈負(fù)相關(guān),其余為正相關(guān)。(3)品種、月齡和組織對(duì)ANGPTL4蛋白的相對(duì)表達(dá)量影響極顯著(P0.001)。品種與月齡的互作對(duì)該蛋白的表達(dá)也有顯著影響(P0.01)。ANGPTL4蛋白的表達(dá)與其mRNA的表達(dá)總體趨勢(shì)一致。(4)檢測(cè)到ANGPTL4基因5'非編碼區(qū)的8個(gè)SNPs中3個(gè)為顛換,5個(gè)為轉(zhuǎn)換。顯著的品種間差異發(fā)生在-1691 CG和-577 TG中。-2361 AG,-2288 GC和-1855AG的等位基因頻率品種間差異不顯著,但在廣靈大尾羊中檢測(cè)到3種基因型,而在小尾寒羊中只檢測(cè)到兩種基因型。-1691 CG和-1625 GA均以正向超顯性遺傳方式影響兩個(gè)品種的尾長、宰前體重和胴體重,以負(fù)向超顯性效應(yīng)方式影響廣靈大尾羊的屠宰率。
[Abstract]:Angiopoietin-like protein 4 (ANGPTL4) promotes both lipid deposition and fatty acid oxidation. On the one hand, it inhibits lipid hydrolysis by inhibiting lipoprotein lipase activity, leading to lipid metabolism-related diseases. On the other hand, it promotes lipolysis by enhancing the activity of catecholamines or glucocorticoids. The structure and function of ANGPTL4 gene, the expression of mRNA and protein, and the polymorphism of the 5'non-coding region of ANGPTL4 gene were studied at three levels of DNA, RNA and protein, and the correlation between the polymorphism and the related traits was analyzed. The coding sequence (CDS) of sheep ANGPTL4 gene was analyzed by bioinformatics. 96 fat-tailed sheep with significant fat-tail difference, Guangling large-tailed sheep and Small-tailed Han sheep, were selected as the study objects. Quantitative real-time PCR (Quantitative real-time PCR) was used. The relative expression levels of ANGPTL4 mRNA in 7 adipose tissues (omentum, omentum, subcutaneous, retroperitoneal,'perirenal, mesenteric and caudal) of 6 age groups were detected and correlated with tail traits and slaughter traits of two breeds. The ANGPTL4 protein in two breeds of ewes was detected by Western blotting. Relative expression levels in adipose tissue (subcutaneous, retroperitoneal, perirenal, and caudal) were measured by PCR-SSCP. Single nucleotide polymorphisms (SNPs) in the 5'-noncoding region of the ANGPTL4 gene were detected in 96 individuals and correlated with related traits. The main results were as follows: (1) The CDS region of the sheep ANGPTL4 gene was 1233 BP long, encoding 410 amino acids. As a secretory protein, it contains a signal peptide with a length of 24 AA and three transmembranes. The amino acid sequence contains one N-glycosylation site, one O-glycosylation site and 18 phosphorylation sites. The results showed that the relationship between sheep and cattle was the closest. (2) The expression of ANGPTL4 mRNA in Guangling large-tailed sheep and Small-tailed Han sheep was different in time and space. The expression of ANGPTL4 mRNA in superficial adipose tissue was higher than that in deep adipose tissue at the age of 0.01. The correlation between the expression of ANGPTL4 mRNA and slaughter and tail type traits was also affected by the interaction between breed and age (P 0.001), breed and tissue (P 0.01) and sex and tissue (P 0.01). The expression of omentum and retroperitoneal fat was negatively correlated with most traits, and the other traits were positively correlated with breed, age and group. The expression of ANGPTL4 protein was also significantly affected by the interaction between varieties and ages (P 0.01). The overall trend of expression of ANGPTL4 protein was consistent with that of its mRNA. (4) Three of the eight SNPs in the 5'non-coding region of ANGPTL4 gene were found to be transposed and five were transposed. Allele frequencies of - 2361 AG, - 2288 GC and - 1855 AG were not significantly different among breeds, but three genotypes were detected in Guangling large tail sheep and only two genotypes were detected in small tail Han sheep. - 1691 CG and - 1625 GA all affected tail length, pre-slaughter weight and carcass by positive superdominant inheritance. Body weight affected the slaughter rate of Guangling large tail sheep by negative overdominance effect.
【學(xué)位授予單位】：山西農(nóng)業(yè)大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2015
【分類號(hào)】：S826;Q78
本文編號(hào)：2230469
[Abstract]:Angiopoietin-like protein 4 (ANGPTL4) promotes both lipid deposition and fatty acid oxidation. On the one hand, it inhibits lipid hydrolysis by inhibiting lipoprotein lipase activity, leading to lipid metabolism-related diseases. On the other hand, it promotes lipolysis by enhancing the activity of catecholamines or glucocorticoids. The structure and function of ANGPTL4 gene, the expression of mRNA and protein, and the polymorphism of the 5'non-coding region of ANGPTL4 gene were studied at three levels of DNA, RNA and protein, and the correlation between the polymorphism and the related traits was analyzed. The coding sequence (CDS) of sheep ANGPTL4 gene was analyzed by bioinformatics. 96 fat-tailed sheep with significant fat-tail difference, Guangling large-tailed sheep and Small-tailed Han sheep, were selected as the study objects. Quantitative real-time PCR (Quantitative real-time PCR) was used. The relative expression levels of ANGPTL4 mRNA in 7 adipose tissues (omentum, omentum, subcutaneous, retroperitoneal,'perirenal, mesenteric and caudal) of 6 age groups were detected and correlated with tail traits and slaughter traits of two breeds. The ANGPTL4 protein in two breeds of ewes was detected by Western blotting. Relative expression levels in adipose tissue (subcutaneous, retroperitoneal, perirenal, and caudal) were measured by PCR-SSCP. Single nucleotide polymorphisms (SNPs) in the 5'-noncoding region of the ANGPTL4 gene were detected in 96 individuals and correlated with related traits. The main results were as follows: (1) The CDS region of the sheep ANGPTL4 gene was 1233 BP long, encoding 410 amino acids. As a secretory protein, it contains a signal peptide with a length of 24 AA and three transmembranes. The amino acid sequence contains one N-glycosylation site, one O-glycosylation site and 18 phosphorylation sites. The results showed that the relationship between sheep and cattle was the closest. (2) The expression of ANGPTL4 mRNA in Guangling large-tailed sheep and Small-tailed Han sheep was different in time and space. The expression of ANGPTL4 mRNA in superficial adipose tissue was higher than that in deep adipose tissue at the age of 0.01. The correlation between the expression of ANGPTL4 mRNA and slaughter and tail type traits was also affected by the interaction between breed and age (P 0.001), breed and tissue (P 0.01) and sex and tissue (P 0.01). The expression of omentum and retroperitoneal fat was negatively correlated with most traits, and the other traits were positively correlated with breed, age and group. The expression of ANGPTL4 protein was also significantly affected by the interaction between varieties and ages (P 0.01). The overall trend of expression of ANGPTL4 protein was consistent with that of its mRNA. (4) Three of the eight SNPs in the 5'non-coding region of ANGPTL4 gene were found to be transposed and five were transposed. Allele frequencies of - 2361 AG, - 2288 GC and - 1855 AG were not significantly different among breeds, but three genotypes were detected in Guangling large tail sheep and only two genotypes were detected in small tail Han sheep. - 1691 CG and - 1625 GA all affected tail length, pre-slaughter weight and carcass by positive superdominant inheritance. Body weight affected the slaughter rate of Guangling large tail sheep by negative overdominance effect.
【學(xué)位授予單位】：山西農(nóng)業(yè)大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2015
【分類號(hào)】：S826;Q78
【參考文獻(xiàn)】
相關(guān)期刊論文 前4條
1 李芬;周佩;楊婷;陳寧博;馬云;;牛ANGPTL2基因結(jié)構(gòu)與功能的生物信息學(xué)分析[J];華北農(nóng)學(xué)報(bào);2012年01期
2 馬云;駱瑩瑩;李芬;王軍芳;徐源;姚瑾;李榮榮;陳宏;;牛ANGPTL4基因結(jié)構(gòu)和功能的生物信息學(xué)分析[J];江西農(nóng)業(yè)大學(xué)學(xué)報(bào);2010年01期
3 馬云;李榮榮;白芳;黃玉鳳;李亞芳;楊婷;陳宏;;黃牛與牦牛ANGPTL4 exon5多態(tài)性與生長性狀的相關(guān)性[J];西北農(nóng)業(yè)學(xué)報(bào);2012年01期
4 朱洪新,李錦軍,覃文新,楊艷華,何祥火,萬大方,顧健人;新基因ANGPTL4的克隆及其在血管新生中的功能研究[J];中華醫(yī)學(xué)雜志;2002年02期
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