基于RNA-Seq技术筛选影响猪肌纤维性状的候选基因

doi:10.11843/j.issn.0366-6964.2019.05.003

摘要/Abstract

摘要：

旨在探究影响肌纤维性状的候选基因和信号通路。本研究以马身猪和大白猪为研究对象，采用HE染色法分析6月龄马身猪和大白猪背最长肌肌纤维直径和密度，利用RNA-Seq分析马身猪和大白猪背最长肌组织中基因表达情况，并对差异表达基因进行GO和KEGG富集分析，再通过qRT-PCR验证RNA-Seq结果的准确性。结果显示，马身猪背最长肌肌纤维直径极显著低于大白猪（P<0.01），而肌纤维密度极显著高于大白猪（P<0.01）。转录组测序结果显示，在6月龄马身猪和大白猪背最长肌中，差异倍数在2倍以上的基因共105个，其中，马身猪相对于大白猪上调的基因有55个，下调的基因有50个。GO富集分析表明，差异表达基因主要富集在与线粒体相关的细胞组分和骨骼肌分化有关的生物学过程中；KEGG分析结果显示，这些差异表达基因主要参与到与氧化磷酸化有关的信号途径。qRT-PCR和RNA-Seq对6个DEGs表达情况的检测结果表明它们的表达趋势相同，说明RNA-Seq结果准确可靠。结合差异基因表达丰度、GO和KEGG富集分析结果，本研究发现，MYL3、MYH3、MYH6基因通过影响肌纤维的组成而影响肌纤维特性，ND6基因通过参与线粒体氧化磷酸化过程影响肌纤维类型，MICU2基因通过维持线粒体Ca²⁺浓度的稳态影响细胞功能，编码转录因子的EGR1和FOS基因通过调节细胞增殖、分化和凋亡等过程影响肌肉生长发育。本研究初步揭示了造成马身猪和大白猪肌纤维性状差异的主要原因，为猪肉品质的改善提供了相应的理论依据。

Abstract:

The aim of this study was to explore the candidate genes and signal pathways affecting muscle fiber characteristics. In this study, muscle fiber diameter and density of longissimus dorsi of Mashen and Large White pigs aged 6 months were measured by HE staining, and the gene expressions in longissimus dorsi of the 2 pig breeds at 6 months old was detected by using RNA-Seq. GO and KEGG enrichment analysis were used to study the function of differentially expressed genes(DEGs). The accuracy of RNA-Seq results was verified by qRT-PCR. The results showed that the fiber diameter of longissimus dorsi of Mashen pig was significantly shorter than that of Large White pig (P<0.01), and the fiber density of Mashen pig was greater than that of Large White pig (P<0.01). A total of 105 DEGs at least 2-fold change were identified, including 55 up-regulated genes and 50 down-regulated genes in longissimus dorsi of Mashen pig compared to Large White pig. GO enrichment analysis showed that the DEGs were mainly enriched in the cellular component related to mitochondrion and skeletal muscle differentiation biological processes, and KEGG analysis showed that these DEGs were mainly involved in the signaling pathways related to oxidative phosphorylation. The expression trends of 6 selected DEGs detected by qRT-PCR and RNA-Seq were the same, which indicated that the results of RNA-Seq were accurate and reliable. Based on the results of expression abundance, GO and KEGG analysis of DEGs, the genes of MYL3, MYH3 and MYH6 were found to affect muscle fiber characteristic by influencing the composition of muscle fiber. ND6 gene was related to muscle fiber type by involving in oxidative phosphorylation pathway in mitochondria. MICU2 gene was involved in keeping homeostasis of Ca²⁺ concentration in mitochondria and further affected cellular function. The genes of EGR1 and FOS coding transcription factors might affect muscle growth and development by regulating cell proliferation, differentiation and apoptosis. This study preliminarily explored the reason causing significantly different muscle fiber characteristics between Mashen and Large White pigs, and provided a theoretical basis for improving meat quality in pig production.

中图分类号:

S828.2

成志敏, 张宁芳, 王媛媛, 乐宝玉, 张万锋, 宋鹏康, 郭晓红, 高鹏飞, 蔡春波, 曹果清, 李步高. 基于RNA-Seq技术筛选影响猪肌纤维性状的候选基因[J]. 畜牧兽医学报, 2019, 50(5): 918-929.

CHENG Zhimin, ZHANG Ningfang, WANG Yuanyuan, LE Baoyu, ZHANG Wanfeng, SONG Pengkang, GUO Xiaohong, GAO Pengfei, CAI Chunbo, CAO Guoqing, LI Bugao. Screening of Candidate Genes for Muscle Fiber Characteristics in Pig Using RNA-Seq[J]. ACTA VETERINARIA ET ZOOTECHNICA SINICA, 2019, 50(5): 918-929.

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