饲粮中添加共轭亚油酸对猪肌肉组织miRNA表达谱的影响

doi:10.11843/j.issn.0366-6964.2018.09.012

摘要/Abstract

摘要：

旨在探讨共轭亚油酸（CLA）通过影响猪肌肉miRNA表达调控肌肉代谢的分子机制。本研究选用体重相近的初产荣昌母猪12头，随机分为对照组和CLA组，CLA组母猪饲粮从妊娠初期开始添加1.5% CLA，持续到仔猪28日龄断奶；断奶后分别从对照组和CLA组挑选仔猪各30头，原CLA组仔猪饲粮中继续添加1.5% CLA。待仔猪体重约30 kg时，每组各选择6头进行屠宰，采集背最长肌和腿肌样品。提取RNA后将对照组和CLA组的背肌和腿肌进行建库；通过Solexa高通量测序技术检测CLA对猪肌肉组织miRNA表达谱的影响，利用生物信息学进行差异显著表达miRNA的靶基因预测和功能分析。结果显示：1）背肌和腿肌分别获得了44 869 982条和45 105 806条clean reads，大多数序列长度在20~23 nt。2）背肌和腿肌分别鉴定到306和304个已知miRNAs，有295个miRNAs共表达。3）添加CLA后能改变猪肌肉中miRNA的表达，分析表明，CLA对腿肌miRNA表达的影响要强于背肌；背肌和腿肌中分别发现5个和12个差异显著表达的miRNAs （P <0.05），其中ssc-miR-224在两种组织中都差异表达，因此共有16个差异表达miRNAs。4）对16个差异表达miRNAs进行靶基因预测和KEGG通路富集分析，发现它们的靶基因主要富集于292个通路，其中24个通路显著富集（P <0.05），包括与肌肉代谢密切相关的MAPK信号通路和Notch信号通路。5）利用荧光定量PCR对随机选择的6个差异表达miRNAs进行验证，证实其表达水平和测序结果基本一致。以上结果说明，CLA可能通过影响肌肉miRNA表达，调控重要肌肉代谢通路。

Abstract:

This study was conducted to investigate the molecular mechanisms of muscle metabolism regulated by conjugated linoleic acid (CLA) through affecting the expression of miRNA in porcine muscle. Twelve purebred Rongchang gilts with similar body weight were randomly divided into control and CLA groups. The pregnant gilts in CLA group were fed with 1.5% CLA diet from the beginning of pregnancy to weaning at 28 days old; After weaning, 30 piglets were respectively selected from the control and CLA groups, and the diet of piglets in original CLA group was also added 1.5% CLA. When the body weight of piglets was about 30 kg, 6 piglets from each group were slaughtered to collect the back muscle and leg muscle samples. After RNA extraction, cDNA libraries were constructed by the back muscle and leg muscle samples of the control and CLA groups; The Solexa deep sequencing was performed to analyze the effect of CLA on the miRNA expression profiles in porcine muscle tissue, the target genes prediction and function analysis for differentially expressed miRNA were carried out through the bioinformatics method. The results showed that:1)There were 44 869 982 and 45 105 806 clean reads in the back muscle and leg muscle tissues, respectively, and most of the small RNA sequence were 20-23 nt. 2) 306 and 304 known miRNAs were respectively found in the back muscle and leg muscle tissues, 295 known miRNAs were co-expressed in two tissues. 3) The expression of miRNA in porcine muscle was changed by CLA supplementation and the effect of CLA on miRNA expression in leg muscle was stronger than that in back muscle; 5 miRNAs in the back muscle and 12 miRNAs in the leg muscle were differentially expressed in corresponding CLA treatment (P<0.05), of which only the ssc-miR-224 was differentially expressed in both two tissues, thus 16 differentially expressed miRNAs were identified. 4) KEGG analysis revealed that the target genes of 16 differentially expressed miRNAs were involved in 292 biological pathways, of which 24 pathways were significantly enriched (P<0.05), including the MAPK and Notch signaling pathways, which were known to regulate muscle metabolism. 5) Six differentially expressed miRNAs were randomly selected to validate the sequencing results by qRT-PCR, the qRT-PCR expression results corresponded well with the results from the sequencing. This study suggests that CLA may regulate muscle metabolic pathways by affecting the expression of miRNA.

中图分类号:

S828.5

王琪, 齐仁立, 刘虹, 王敬, 黄金秀. 饲粮中添加共轭亚油酸对猪肌肉组织miRNA表达谱的影响[J]. 畜牧兽医学报, 2018, 49(9): 1908-1918.

WANG Qi, QI Ren-li, LIU Hong, WANG Jing, HUANG Jin-xiu. Effects of Conjugated Linoleic Acid Supplementation in Diet on the Expression Profile of miRNAs in Porcine Muscle Tissue[J]. ACTA VETERINARIA ET ZOOTECHNICA SINICA, 2018, 49(9): 1908-1918.

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