基于血液转录组的林麝免疫相关基因表达的研究

Animal immune defense is a complex network system, which involved a large number of immune related genes and different immune pathways and their interaction. The traditional method is difficult to accurately assess the interaction of immune related genes, whereas blood transcriptome analysis can accurately describe the immune related genes and their regulation pathways, and explain the basic mechanisms of disease progression. Based on our previous research, we will sequence and map the blood transcriptomes of different age and gender groups of abscessed and healthy Forest musk deer (Moschus berezovskii) by using Illumina HiSeq 2500 high-throughput sequencing technology. We will analyze the changes of gene expression in the different groups of Forest musk deer and detect differentially expressed genes. The influence of age, gender, and abscess on the gene expression will be explored after assembling the blood transcriptomes. And we will also analyze expression features of immune related genes and regulation of immune pathways in the abscess process of Forest musk deer. Furthermore, we will identify the distribution characteristics of SNP (Single nucleotide polymorphism) in the blood transcriptomes of different age and gender groups of abscessed and healthy Forest musk deer. The SNPs associated with immune genes are searched in the transcriptome level of Forest musk deer. Based on the previous genome study, We will explore the relationship between SNPs and expression level of immune related genes, and seek for the molecular markers of disease detection. The research findings will provide the scientific evidence and new idea in order to health management, disease diagnosis and therapy of captive Forest musk deer.

动物免疫防御是复杂的网络系统，涉及大量的免疫相关基因和不同的免疫通路及其相互作用，传统方法难以准确评估免疫相关基因的相互作用，高通量血液转录组分析可精确地描述免疫相关基因及其调控通路，阐释疾病发生的机制。因此本研究拟在前期研究基础上，采用Illumina HiSeq 2500高通量测序技术对不同年龄、不同性别患脓肿病和健康林麝的血液进行转录组测序，分析其基因表达水平的变化，寻找差异表达基因，探索年龄、性别和脓肿病对其免疫及其相关基因表达的影响，并分析林麝在脓肿病发生过程中免疫通路的变化及相关基因的表达特性。另外，鉴定不同年龄、不同性别患脓肿病和健康林麝转录组之间SNP（单核苷酸多态性）位点的分布特性，并结合前期林麝基因组的研究，在转录组水平鉴定与免疫基因相关的SNP位点，探索其与免疫相关基因表达量的关系，寻找林麝疾病检测的分子标记，为林麝健康管理和疾病的诊断和治疗提供科学依据。

林麝（Moschus berezovskii）是国家一级重点保护野生动物，麝资源面临枯竭。在本研究中，我们完成了36只不同年龄段和不同性别的脓肿病和健康林麝的血液转录组测序。在脓肿病和健康林麝之间筛选到300个差异表达基因具有显著统计学意义，其中上调基因有243个，下调基因有57个。298个差异表达基因有基因注释，其中包括242个上调基因和56个下调基因。GO功能富集分析表明，最显著性富集的10个生物过程GO条目中，大多数与免疫相关，比如炎症反应、免疫应答和防御反应，暗示大多数差异表达基因属于免疫相关基因。KEGG通路富集分析显示差异表达基因富集到15个通路上，其中，蛋白酶体通路是最显著性富集的免疫通路。初步表明年龄和脓肿病对其免疫相关基因表达有一定影响，而患脓肿病和健康林麝SNP对基因表达没有显著影响。另外，在脓肿病和健康林麝之间筛选到267个差异表达miRNA具有显著统计学意义，其中上调miRNAs有153个，下调miRNAs有114个。193个差异表达的miRNAs有注释，其中包括115个上调miRNAs和78个下调miRNAs。初步鉴定出138个免疫相关的miRNAs，其中有50个保守的miRNAs。在脓肿病和健康林麝之间共识别和鉴定19,880个lncRNAs，寻找到1301个差异表达miRNA具有显著统计学意义，其中有467个显著上调lncRNAs，有834个显著下调的lncRNAs。功能富集分析表明这些差异表达lncRNAs主要与免疫相关。我们成功筛选出21个多态性高、稳定性好、特异性强的分子标记，将有助于为疾病的诊断和林麝健康管理提供科学依据。