本文關(guān)鍵詞： CLA 榮昌豬 肉品質(zhì) miRNA　出處：《西南大學(xué)》2017年碩士論文　論文類型：學(xué)位論文

【摘要】：本論文研究了日糧長期添加共軛亞油酸(Conjugated linoleic acids,CLA)對榮昌豬肉品質(zhì)及相關(guān)基因表達(dá)的影響,并通過高通量microRNA(miRNA)測序和生物信息學(xué)分析,深入探究了CLA對豬肌肉miRNAs表達(dá)的影響,為CLA在畜禽養(yǎng)殖中的廣泛應(yīng)用提供理論依據(jù)和試驗(yàn)參考。試驗(yàn)選取50頭日齡和體重相近的健康榮昌仔豬(公母各半),采用單因素處理安排,隨機(jī)分為2組:對照組和CLA組,每組5個(gè)重復(fù),每個(gè)重復(fù)5頭豬(組間體重?zé)o顯著差異)。從仔豬斷奶開始持續(xù)在CLA組日糧中添加1.5%的CLA混合物,等量替換大豆油。對照組與CLA組的日糧營養(yǎng)水平相同。當(dāng)試驗(yàn)豬的平均體重約達(dá)85kg時(shí),分別從每個(gè)重復(fù)隨機(jī)選出1頭進(jìn)行稱重、屠宰和肉質(zhì)指標(biāo)測定;同時(shí)收集背最長肌樣品用于肉質(zhì)相關(guān)基因表達(dá)測定和mi RNA高通量測序。主要研究結(jié)果如下:1、日糧添加CLA對榮昌豬肉質(zhì)及相關(guān)基因表達(dá)的影響(1)與對照組相比,CLA組的平均背膘厚下降了7.55%;肌內(nèi)脂肪含量提高了24.71%(P0.1);肉色評分提高了9.3%(P0.1),肉色b*值顯著降低了22.7%(P0.05);肌纖維密度提高了23.47%;同時(shí),肌肉中T-AOC活性顯著提高(P0.05),MDA含量極顯著減少(P0.01)。(2)與對照組相比,CLA組豬肌肉中脂肪沉積調(diào)控因子PPARγ、PGC-1α、FABP4和FAS基因表達(dá)顯著升高(P0.05),C/EBPα和FATP1基因表達(dá)極顯著升高(P0.01);肌纖維MyHC1基因表達(dá)顯著升高(P0.05),MyHC2a基因表達(dá)極顯著升高(P0.01)。2、日糧添加CLA對榮昌豬肌肉miRNAs表達(dá)的影響(1)構(gòu)建對照組和CLA組small RNA文庫,經(jīng)Solexa測序并過濾,得到的clean reads比對到mi RBase21.0數(shù)據(jù)庫和豬基因組數(shù)據(jù)庫。兩個(gè)RNA文庫中共鑒定出556個(gè)miRNAs,其中有472個(gè)miRNAs共表達(dá)。與對照組相比,CLA組文庫中有96個(gè)miRNAs表達(dá)極顯著下調(diào)(P0.01),156個(gè)miRNAs表達(dá)極顯著上調(diào)(P0.01)。(2)13個(gè)miRNAs受CLA的影響表達(dá)發(fā)生極顯著改變(P0.01),且拷貝數(shù)大于500,上調(diào)倍數(shù)大于1.5或下調(diào)倍數(shù)小于0.6,其中7個(gè)(miRNA-1、miRNA-206、miRNA-335、miRNA-486、miRNA-133b、miRNA-186、miRNA-374a-3p)參與調(diào)控肌肉生長和發(fā)育,3個(gè)(miRNA-30e、miRNA-21、miRNA-199a-5p)參與調(diào)控脂肪代謝,3個(gè)(miRNA-19b、miRNA-127、miRNA-7)涉及癌癥發(fā)生和發(fā)展。(3)對參與調(diào)控肌肉生長和發(fā)育以及脂肪代謝的miRNAs進(jìn)行靶基因預(yù)測,發(fā)現(xiàn)10個(gè)miRNAs共有2815個(gè)靶基因;對靶基因進(jìn)行KEGG信號通路富集分析,發(fā)現(xiàn)主要顯著富集在代謝通路(ssc01100:Metabolic pathways,P0.01)和AMPK信號通路(ssc04152:AMPK signaling pathway,P0.01)等。試驗(yàn)所觀測的脂肪代謝因子如PPARγ、PGC-1α和FAS等都處于AMPK信號通路下游,提示CLA可能通過miRNAs作用于AMPK信號通路,調(diào)控下游脂肪代謝基因,進(jìn)而影響豬的肌肉脂肪代謝和肉質(zhì)性狀。綜上所述,日糧中長期添加適量CLA能在一定程度上改善豬肉品質(zhì),并顯著改變肌肉中肉質(zhì)相關(guān)基因和miRNAs表達(dá),但CLA通過miRNAs改善豬肉品質(zhì)的具體作用機(jī)制仍需進(jìn)一步探討。
[Abstract]:The effects of conjugated linoleic acid (CLA) supplementation on the quality of Rongchang pork and the expression of related genes were studied in this paper. The effects of CLA on the expression of miRNAs in pig muscle were studied by high throughput microRNAs sequencing and bioinformatics analysis. To provide theoretical basis and experimental reference for the wide application of CLA in livestock and poultry breeding, 50 healthy Rongchang piglets of similar body weight and day old (male and female) were selected and randomly divided into two groups: control group and CLA group. There was no significant difference in body weight among the five pigs in each group (no significant difference in body weight). From weaning piglets, 1.5% CLA mixture was added to the diet of CLA group. The same amount of soybean oil was replaced. When the average weight of the pigs in the control group and the CLA group was about 85 kg, one pig was selected randomly from each repeat for weighing, slaughtering and meat quality indexes. At the same time, the longissimus dorsi muscle samples were collected for meat quality related gene expression determination and mi RNA high throughput sequencing. The main results were as follows: 1. The effect of dietary CLA supplementation on the expression of pork quality and related genes in Rongchang. The average backfat thickness decreased 7.55%; the intramuscular fat content increased 24.71% P0.1; the meat color score increased 9.3% P0.1; the meat color b* value significantly decreased 22.7% P0.05; the muscle fiber density increased 23.47%; at the same time, Compared with the control group, the expression of PPAR 緯 -PGC-1 偽 -FABP4 and FAS gene in muscle was significantly increased, and the expression of C- EBP 偽 and FATP1 gene in muscle was significantly higher than that in the control group, and the expression of MyHC1 gene in muscle fiber was significantly higher than that in the control group. The expression of MyHC2a gene was significantly increased in Rong Chang pigs. The effect of dietary CLA supplementation on the expression of miRNAs in Rong Chang pig muscle was significantly increased. The small RNA library of control group and CLA group was constructed. After Solexa sequencing and filtering, The obtained clean reads was compared to the mi RBase21.0 database and the porcine genomic database. 556 miRNAss were identified from the two RNA libraries, of which 472 were coexpressed. Compared with the control group, 96 of the RNA libraries had significantly down-regulated miRNAs expression and 156 miRNAs. The expression of 13 miRNAs was significantly up-regulated by CLA, and the number of copies was greater than 500, the upregulation multiple was greater than 1.5 or the down-regulation multiple was less than 0.6. Seven miRNA-1 miRNA-335mRNA-486miRNA-133bmiRNA-186miRNA-374a-3p) were involved in the regulation of muscle growth and development. 3 miRNA-30esimi-21RNA RNAmiRNAmiRNA-199a-5p) were involved in the regulation of muscle growth and development. Fat metabolism, 3 miRNA-19bsimiRNA-127 miRNA-7) involved in carcinogenesis and development. 3) target gene prediction of miRNAs involved in regulating muscle growth and development as well as fat metabolism. A total of 2 815 target genes were found in 10 miRNAs, and the KEGG signaling pathway enrichment analysis was carried out on the target genes. It was found that it was mainly enriched in the metabolic pathway ssc0100: Metabolic pathways0.01) and the AMPK signaling pathway ssc04152: AMPK signaling pathway P0.01). The observed fatty metabolic factors such as PPAR 緯 PGC-1 偽 and FAS were located downstream of the AMPK signaling pathway, suggesting that CLA may act on AMPK signaling pathway via miRNAs. In conclusion, medium and long term dietary CLA supplementation could improve pork quality and significantly change the expression of fleshy related genes and miRNAs in the muscle. However, the mechanism of CLA improving pork quality through miRNAs still needs to be further explored.
【學(xué)位授予單位】：西南大學(xué)
【學(xué)位級別】：碩士
【學(xué)位授予年份】：2017
【分類號】：S828

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本文編號：1500406