两品种猪快慢肌纤维差异表达基因鉴定与通路分析

发布时间：2018-03-14 06:39

本文选题：猪　切入点：快慢肌　出处：《华中农业大学》2015年硕士论文　论文类型：学位论文

【摘要】：猪肉是我国肉类蛋白质食物的主要来源之一,养猪业在我国农业中占有举足轻重的地位。肉质受到肌纤维性状的影响,其中,肌纤维类型被认为是影响肉质的重要因素之一。遗传背景及人工选育方式的不同,造成了中外猪种在生产性状方面存在很大的差异。陆川猪作为我国特有的猪种资源,具有肉质优良的种质特性;大白猪作为西方猪种的代表性品种,虽然在生长速度方面具有优势,但是其相对较差的肉品质已不满足于消费者的需求。通过高通量测序的技术来分析品种间/不同特性的肌肉间的差异是一种高效科学的方法,对转录组的分析,可以让我们知道究竟是哪些基因或者通路造成了这些性状上的差异。本研究通过对品种间以及快慢肌间进行了表型和转录组差异的分析,初步探索了产生差异的原因,为培育出符合畜牧业发展需求的优秀猪品种提供了素材。主要研究结果如下:1、运用石蜡切片、HE染色、免疫组织化学技术,对陆川猪和大白猪的比目鱼肌、趾长伸肌、股二头肌和背最长肌进行了肌纤维表型的试验与分析。结果显示,在这四块肌肉中陆川猪肌纤维直径均显著小于大白猪。在品种间的比较中,比目鱼肌和背最长肌中慢肌纤维所占比例,陆川猪要显著低于大白猪,但是趾长伸肌和股二头肌中慢肌纤维比例,陆川猪却要显著高于大白猪;在陆川猪上,慢肌纤维的比例:股二头肌比目鱼肌趾长伸肌背最长肌;大白猪上,慢肌纤维的比例:比目鱼肌背最长肌股二头肌趾长伸肌。2、运用高通量转录组测序技术,成功对陆川猪和大白猪的比目鱼肌和趾长伸肌进行了RNA-Seq建库测序,数据质检合格后对基因的表达水平进行了分析。结果显示,各分组间基因整体表达量基本相同;在所有样本中,大约有超过24,500个基因表达,其中有16,404个基因在所有样本中表达;品种特异性表达的基因,陆川猪中有825个,大白猪有366个,而在两品种内均表达的基因有17,248个;所有样本中,转录起始位点(TSS)剪切、转录终止位点(TTS)剪切是两种分布最多的剪切类型,在所有样本中约占总可变剪切事件的80%。3、对各比较组进行了差异表达基因的筛选,最终得到了的差异表达基因中有754个已知基因;在这些基因中,有229个基因在品种间两个比较组中均差异表达;有14个基因在肌肉间两个比较组中均差异表达,为三个比较组所共有的基因共有20个,已知基因有17个。4、对筛选出的差异表达基因以比较组为单位,分别进行GO分析,分别有59(YSOL vs YEDL)、32(LEDL vs YEDL)和77(LSOL vs YSOL)个GO term被富集。这些被显著富集到的GO term包括一些与肌肉骨骼生长发育(muscle organ development、skeletal system development、skeletal muscle tissue development)、能量代谢相关(gluconeogenesis、lipid storage、monosaccharide metabolic process)和离子转运(metal ion transport、organic anion transport)等生物学过程;hexose metabolic process、glucose metabolic proces、monosaccharide metabolic process、rhythmic process,这四个GO term在三个比较组中均被富集。5、对差异表达基因进行KEGG分析后发现,四个比较组的差异表达基因最终分别显著富集到了16(LSOLvs YSOL)、11(LEDLvs YEDL)、4(LSOLvs LEDL)和9(YSOLvs YEDL)条通路中。被显著富集的通路包括糖酵解(Glycolysis/Gluconeogenesis)通路、三羧酸循环(TCA cycle)和有关脂肪酸代谢的途径(Fatty acid degradation、Fatty acid metabolism);有15个通路是四个比较组所共有,其中包括MAPK信号通路、PI3K-Akt信号通路等。
[Abstract]:Pork is the main source of China's meat protein food, pig industry occupies a pivotal position in China's agriculture. The meat is affected, muscle fiber traits, muscle fiber type is considered to be one of the important factors affecting meat quality. Genetic background and artificial breeding method is different, resulting in the production of Chinese and foreign pig breeds there are great differences between the characters. As Lu Chuan pig pig resource of China, having fleshy good germplasm characteristics; large white pigs as western representative varieties, although has the advantage in the growth rate, but its relatively poor meat quality has not satisfied the demand of consumers. By high-throughput sequencing the technology to analyze the differences between the varieties / different characteristics between muscles is an efficient method of science, analysis of the transcriptome, can let us know exactly what the gene or pathway made The differences in these traits. This study of varieties and analyzed the difference between fast and slow muscle phenotype and transcriptome, explores the reasons for the differences, and provide materials for breeding of animal husbandry in good accord with the development requirements of pig breeds. The main results are as follows: 1, the use of paraffin sections. HE staining, immunohistochemical technique, the soleus muscle of Lu Chuan and large white pigs, extensor digitorum longus, unit two biceps and longissimus muscle of the test and analysis of muscle fiber phenotype. The results showed that in the four muscles in Lu Chuan pig muscle fiber diameter was significantly smaller than in large white pigs. Among the varieties, and soleus muscle in longissimus dorsi muscle of slow muscle fiber proportion was significantly lower than that of large white pig Lu Chuan, but the extensor digitorum longus muscle and the femoral head two slow muscle fiber ratio, Lu Chuan is significantly higher than that of large white pig; in Lu Chuan pig, slow muscle fiber The proportion of femoral head two twitchmuscle extensor digitorum longus longissimus muscle; large white, slow muscle fiber ratio: soleus muscle longissimus muscle unit two biceps extensor digitorum longus.2, using high-throughput transcriptome sequencing technology, the success of Lu Chuan and large white pigs of the soleus and extensor digitorum longus the RNA-Seq library sequencing, data quality inspection on the level of gene expression was analyzed. The results showed that the gene expression of the whole group is basically the same; in all samples, there are about more than 24500 genes, including 16404 genes expressed in all samples; cultivar specific expression gene, Lu Chuan there are 825 pigs, large white pigs 366, which were expressed in two varieties in 17248 genes; all the samples, the transcription initiation site (TSS) shear, transcription termination sites (TTS) are two types of shear shear distribution of the largest, in all samples The total variable shear event 80%.3, the comparison group for the screening of differentially expressed genes, the difference obtained expression of 754 known genes; among these genes, the expression of two groups were compared between the 229 genes in varieties; expression of two groups were compared the difference of 14 genes in muscle between three groups compared the common gene of a total of 20 known genes, 17.4 of the differentially expressed genes in the comparison group as a unit, were analyzed by the GO. There were 59 (YSOL vs YEDL), 32 (LEDL vs YEDL) and 77 (LSOL vs YSOL) GO term was enriched. These were significantly enriched to GO term include some skeletal muscle growth and development (muscle organ development, skeletal system development, skeletal muscle tissue development), energy metabolism (gluconeogenesis, lipid storage, monosaccharide Metab Olic process (metal) and ion transport ion transport, organic anion transport hexose metabolic) and other biological processes; process glucose, metabolic proces, monosaccharide metabolic process, rhythmic process, the four GO term in three groups were enriched in.5, the expression of KEGG gene was found on analysis of differences in genes ultimately respectively. Enrichment to 16 the expression of the four comparison groups (LSOLvs YSOL 11), the difference (LEDLvs YEDL), 4 (LSOLvs LEDL) and 9 (YSOLvs YEDL) pathways were significantly enriched. Pathways including glycolysis (Glycolysis/Gluconeogenesis) pathway, tricarboxylic acid cycle three (TCA cycle) and related to fatty acid metabolism (Fatty acid degradation Fatty way acid metabolism); there are 15 pathways is a comparison of the four groups there, including MAPK signaling pathway, PI3K-Akt signaling pathway.

【学位授予单位】：华中农业大学
【学位级别】：硕士
【学位授予年份】：2015
【分类号】：S828

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