共轭亚油酸对荣昌猪肉质相关基因及肌肉microRNAs表达的影响

发布时间：2018-02-10 11:25

本文关键词： CLA 荣昌猪肉品质 miRNA　出处：《西南大学》2017年硕士论文　论文类型：学位论文

【摘要】：本论文研究了日粮长期添加共轭亚油酸(Conjugated linoleic acids,CLA)对荣昌猪肉品质及相关基因表达的影响,并通过高通量microRNA(miRNA)测序和生物信息学分析,深入探究了CLA对猪肌肉miRNAs表达的影响,为CLA在畜禽养殖中的广泛应用提供理论依据和试验参考。试验选取50头日龄和体重相近的健康荣昌仔猪(公母各半),采用单因素处理安排,随机分为2组:对照组和CLA组,每组5个重复,每个重复5头猪(组间体重无显著差异)。从仔猪断奶开始持续在CLA组日粮中添加1.5%的CLA混合物,等量替换大豆油。对照组与CLA组的日粮营养水平相同。当试验猪的平均体重约达85kg时,分别从每个重复随机选出1头进行称重、屠宰和肉质指标测定;同时收集背最长肌样品用于肉质相关基因表达测定和mi RNA高通量测序。主要研究结果如下:1、日粮添加CLA对荣昌猪肉质及相关基因表达的影响(1)与对照组相比,CLA组的平均背膘厚下降了7.55%;肌内脂肪含量提高了24.71%(P0.1);肉色评分提高了9.3%(P0.1),肉色b*值显著降低了22.7%(P0.05);肌纤维密度提高了23.47%;同时,肌肉中T-AOC活性显著提高(P0.05),MDA含量极显著减少(P0.01)。(2)与对照组相比,CLA组猪肌肉中脂肪沉积调控因子PPARγ、PGC-1α、FABP4和FAS基因表达显著升高(P0.05),C/EBPα和FATP1基因表达极显著升高(P0.01);肌纤维MyHC1基因表达显著升高(P0.05),MyHC2a基因表达极显著升高(P0.01)。2、日粮添加CLA对荣昌猪肌肉miRNAs表达的影响(1)构建对照组和CLA组small RNA文库,经Solexa测序并过滤,得到的clean reads比对到mi RBase21.0数据库和猪基因组数据库。两个RNA文库中共鉴定出556个miRNAs,其中有472个miRNAs共表达。与对照组相比,CLA组文库中有96个miRNAs表达极显著下调(P0.01),156个miRNAs表达极显著上调(P0.01)。(2)13个miRNAs受CLA的影响表达发生极显著改变(P0.01),且拷贝数大于500,上调倍数大于1.5或下调倍数小于0.6,其中7个(miRNA-1、miRNA-206、miRNA-335、miRNA-486、miRNA-133b、miRNA-186、miRNA-374a-3p)参与调控肌肉生长和发育,3个(miRNA-30e、miRNA-21、miRNA-199a-5p)参与调控脂肪代谢,3个(miRNA-19b、miRNA-127、miRNA-7)涉及癌症发生和发展。(3)对参与调控肌肉生长和发育以及脂肪代谢的miRNAs进行靶基因预测,发现10个miRNAs共有2815个靶基因;对靶基因进行KEGG信号通路富集分析,发现主要显著富集在代谢通路(ssc01100:Metabolic pathways,P0.01)和AMPK信号通路(ssc04152:AMPK signaling pathway,P0.01)等。试验所观测的脂肪代谢因子如PPARγ、PGC-1α和FAS等都处于AMPK信号通路下游,提示CLA可能通过miRNAs作用于AMPK信号通路,调控下游脂肪代谢基因,进而影响猪的肌肉脂肪代谢和肉质性状。综上所述,日粮中长期添加适量CLA能在一定程度上改善猪肉品质,并显著改变肌肉中肉质相关基因和miRNAs表达,但CLA通过miRNAs改善猪肉品质的具体作用机制仍需进一步探讨。
[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 纬 -PGC-1 伪 -FABP4 and FAS gene in muscle was significantly increased, and the expression of C- EBP 伪 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 纬 PGC-1 伪 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.
【学位授予单位】：西南大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：S828

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