整合转录组及全基因组重测序方法鉴定影响猪脂肪沉积的关键基因及其变异

发布时间：2018-03-29 11:53

本文选题：猪　切入点：脂肪沉积　出处：《中国农业大学》2015年博士论文

【摘要】：猪的脂肪沉积的多少严重影响着猪肉的生产效率、猪肉品质、繁殖性能以及消费者对猪肉的选择。本研究通过B型超声波背膘仪对松辽黑猪和长白猪群体进行活体背膘厚度的测定,筛选出具有极端背膘厚度的个体。利用高通量测序技术检测筛选出的具有极端背膘厚度的松辽黑猪和长白猪的背部皮下脂肪(各3对)和肝脏(各2对)的转录组,筛选出差异表达基因,确定在脂肪组织和肝脏中调控脂肪沉积的关键基因。同时,针对每个品种不同极端厚度的群体(高、低)的基因组进行混池重测序,找到2组之间的基因变异。通过整合转录组数据、重测序数据及QTL数据库等以及功能基因组学、生物信息学等方法,分析差异表达基因、基因变异的变化规律及相互关系,筛选、鉴定并验证影响猪脂肪沉积的候选基因及其变异。本研究对比了不同的品种及不同表型个体间的肉品质。松辽黑猪与长白猪的肉色之间有着显著的差异,同时又有着更高的肌内脂肪含量和更低失水率。而在同一品种的不同背膘厚度的群体之间,只有肌内脂肪含量有极显著的差异,其他肉品质之间并没有显示出差异。通过转录组测序,在具有极端背膘厚度的松辽黑猪的脂肪组织和肝脏中,分别找到了188个和91个差异表达基因。在长白猪的脂肪组织和肝脏中,也分别找到了552个和72个差异表达基因。通过生物学功能分析发现,差异表达基因参与了脂肪酸合成、脂肪合成底物的合成、脂类代谢的激活、胰岛素信号通路、PPAR信号通路、MAPK信号通路、PPAR信号通路等众多与脂肪沉积相关的生物学过程。这些与脂肪合成、代谢相关的基因如1ASN、PCK1、ME1、ELOVL6可以被认为影响脂肪沉积多少的候选基因。同时,在脂肪组织中找到了大量的免疫相关的差异表达基因,结果表明脂肪组织作为一个免疫器官,脂肪沉积的多少直接影响着脂肪组织在免疫中的作用。通过全基因组重测序中,在整个基因组中找到了大量基因变异,包括SNP和插入缺失,这些基因变异大部分坐落在基因间或内含子上。同时针对含有可能影响基因表达和功能的错义突变、移码突变的基因进行功能富集分析,发现在脂肪合成、转运、储存、代谢等通路上含有大量的遗传变异。大量位点的发现为进一步研究影响脂肪沉积提供了基础。最后整合转录组测序和全基因组重测序的结果,找到关键差异表达基因上的变异。在脂肪酸合成的关键基因FASN、ACACA、ME1、PCK1等上找到很多的变异,特别是存在这些基因的错义突变、UTR突变等,都是可能影响脂肪沉积的重点突变。本研究发现了脂肪合成关键的基因如ELOVL6、LDHA、CYP2B224坐落在脂肪沉积相关的QTL上。本研究同时挑选出了部分变异位点,与背膘厚度进行关联分析,发现ME1、PCK1基因UTR区上变异位点显著影响背膘的厚度。综上所述,脂肪组织中的脂肪酸合成能力是影响背膘厚度的关键因素,同时肝脏和脂肪组织中的代谢也发挥着重要的作用。研究结果全面阐述了不同背膘厚度的松辽黑猪和长白猪脂肪组织和肝脏基因表达谱及基因组的变异,为深入研究猪脂肪沉积的分子调控机制及进一步的实验奠定了基础。
[Abstract]:How much of the fat deposition of pigs seriously affects the pork production efficiency, pork quality, reproduction performance and consumer choice of pork. In this study, determination of backfat thickness on Songliao Black Pig and Landrace populations by B type ultrasonic backfat instrument, and screened with extreme backfat thickness individuals screened by. High throughput sequencing technology has the extreme backfat thickness of Songliao Black Pig and Landrace back fat (3 pairs) and liver (2 pairs) transcriptome, we screened the differentially expressed genes, determine the key genes regulating control of fat deposition in adipose tissue and liver. At the same time, for each of the different varieties of extreme the thickness of the groups (high, low) of the genome of the mixed pool re sequencing, find the genetic variation between the 2 groups. By integrating transcriptome data, re sequencing data and QTL database and functional genomics, bioinformatics In order to identify and verify candidate genes and their variation affecting porcine fat deposition, we analyzed the change rules and relationship of differentially expressed genes and gene mutation.
This study compared the different varieties and different phenotypes between individuals of meat quality. There are significant differences between Songliao Black Pig and Landrace flesh, and has a fat content and lower water loss rate is higher in the muscle. And in the same variety of different backfat thickness between the groups, only the fat content there were significant differences in muscle, and did not show differences between other meat quality.
Through transcriptome sequencing, in adipose tissue and liver with extreme backfat thickness in Songliao Black pig, were found 188 and 91 differentially expressed genes in adipose tissue and liver in Landrace pigs, were also found 552 and 72 differentially expressed genes. The analysis can be found through biological function, difference the expression of genes involved in fatty acid synthesis, the synthesis of fat synthesis substrate activation, lipid metabolism, insulin signaling pathway, PPAR signaling pathway, MAPK signaling pathway, PPAR signaling pathway and many other biological processes associated with fat deposition. These fat synthesis, metabolism related genes such as 1ASN, PCK1, ME1, ELOVL6 can be considered how many of the candidate gene affecting fat deposition in adipose tissue. At the same time, to find a large number of immune related gene expression. The results suggest that adipose tissue as an immune organ, how much fat deposition It has a direct effect on the role of adipose tissue in the immune system.
Through whole genome sequencing, to find a large number of mutations in the genome, including SNP and indel, most of these mutations located in genes or introns. At the same time for containing may affect missense gene expression and function of gene mutation, mutation of shift function enrichment analysis, found in fat synthesis, transport, storage, metabolic pathway containing a large amount of genetic variation. A large number of sites that provide a basis for further study on the effects of fat deposition.
Finally, the integration of transcriptome sequencing and whole genome sequencing results, find the key differences between the expression of the mutations in key genes in fatty acid synthesis. FASN, ACACA, ME1, PCK1 and so on to find a lot of variation, especially in the presence of missense mutations in these genes, UTR mutation, are likely to influence on fat deposition mutation. This study found a fat synthesis of key genes such as ELOVL6, LDHA, CYP2B224 is located in QTL related to fat deposition. This study also selected some mutation sites for association analysis, found that ME1 and backfat thickness, ectopic point significantly affect backfat thickness PCK1 gene UTR region.
In summary, fatty acid synthesis ability in adipose tissue is a key factor affecting the backfat thickness at the same time, liver and adipose tissue metabolism also plays an important role. The research results comprehensively expounds the variation of Songliao Black Pigs and Landrace pigs adipose tissue and liver gene expression profiles of different backfat thickness and genome, laid the foundation molecular regulation mechanism research of pig fat deposition and further experiments.

【学位授予单位】：中国农业大学
【学位级别】：博士
【学位授予年份】：2015
【分类号】：S828

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