貉外周血单核细胞转录组RNA-Seq数据的de novo拼接和信息比对研究

De novo splicing and information study of PBMCs RNA-Seq transcription of raccoon dog

为从转录组水平上分析貉免疫分子相关信息,深入了解并研究其免疫防御机制,本研究采用2代测序技术,对貉外周血单核细胞(PBMCs)进行了RNA-Seq测序、de novo拼接和信息比对研究。结果表明,转录组测序共得到3.1 GB的原始数据,共32 245 804个读长,去除载体信息的数据量为28 797 350个读长。经质量控制和de novo拼接后,对组装的转录本利用Trans Decoder鉴定编码区域,共获得118 868条貉转录本,其平均长度为525.53 nt。利用Trinotate对ORF和contigs进行功能注释,利用Uniprot database、RNAMMER、egg NOG、GO、KEGG对预测的序列进行注释。结果显示,COG功能注释中信号转导机制(T)及防御机理(V),KEGG注释通路中B/T-细胞受体信号通路、TLR/NLR/RIR受体信号通路、吞噬体通路、自然杀伤细胞介导的细胞毒性通路均与貉免疫应答和抗病相关。本研究为后续开展貉进化研究及免疫机制研究提供参考,为貉全基因组测序项目提供较好的注释基础。

In order to study the immunologic mechanism of raccoon dog, the next generation sequencing technology was implemented using RNA sequencing, de novo splicing, and information comparison of the peripheral blood mononuclear cells transcriptome. The results showed that the transcriptome sequencing had received a total of 3.1 GB raw data and 32 245 804 reads. The number of reads after removing the information of carriers was 28 797 350. Using the TransDecoder to identify the coding regions of the assembled transcripts after the quality controlling and the de novo splicing, there was a number of 118 868 raccoon dog transcripts with the average length of 525.53 nt. Furthermore, functional annotation of ORF and contigs using Trinotate, and prediction of sequence information using Uniprot database, RNAMMER, eggNOG, GO, KEGG showed that, B cell or T cell receptor signaling pathway, Toll-like, NOD-like or RIG-like receptor signaling pathway, Phagosome pathways and Cytotoxic pathways mediated by NKT cell of KEGG annotation, as well as signal transduction mechanisms and defense mechanisms of COG annotation involved in immune response and disease-resistant activities of raccoon dog. The study provides a foundation and reference for further study of evolution, immunologic mechanism, and whole genome sequencing of raccoon dog.