Table_6_Full-length transcriptome sequencing and comparative transcriptome analysis of Eriocheir sinensis in response to infection by the microsporidian Hepatospora eriocheir.xlsx

As a new generation of high-throughput sequencing technology, PacBio Iso-Seq technology (Iso-Seq) provides a better alternative sequencing method for the acquisition of full-length unigenes. In this study, a total of 22.27 gigabyte (Gb) subread bases and 128,614 non-redundant unigenes (mean length: 2,324 bp) were obtained from six main tissues of Eriocheir sinensis including the heart, nerve, intestine, muscle, gills and hepatopancreas. In addition, 74,732 unigenes were mapped to at least one of the following databases: Non-Redundant Protein Sequence Database (NR), Gene Ontology (GO), Kyoto Encyclopaedia of Genes and Genomes (KEGG), KEGG Orthology (KO) and Protein family (Pfam). In addition, 6696 transcription factors (TFs), 28,458 long non-coding RNAs (lncRNAs) and 94,230 mRNA-miRNA pairs were identified. Hepatospora eriocheir is the primary pathogen of E. sinensis and can cause hepatopancreatic necrosis disease (HPND); the intestine is the main target tissue. Here, we attempted to identify the key genes related to H. eriocheir infection in the intestines of E. sinensis. By combining Iso-Seq and Illumina RNA-seq analysis, we identified a total of 12,708 differentially expressed unigenes (DEUs; 6,696 upregulated and 6,012 downregulated) in the crab intestine following infection with H. eriocheir. Based on the biological analysis of these DEUs, several key processes were identified, including energy metabolism-related pathways, cell apoptosis and innate immune-related pathways. Twelve selected genes from these DEUs were subsequently verified by quantitative real-time PCR (qRT-PCR) analysis. Our findings enhance our understanding of the E. sinensis transcriptome and the specific association between E. sinensis and H. eriocheir infection.

作为新一代高通量测序技术，PacBio Iso-Seq测序技术（Iso-Seq）为全长单基因簇（unigene）的获取提供了更优质的测序方案。本研究从中华绒螯蟹（Eriocheir sinensis）的六大主要组织——心脏、神经组织、肠道、肌肉、鳃及肝胰腺中，共获得22.27吉字节（Gb）的子读段碱基，组装得到128,614个非冗余单基因簇，其平均长度为2324 bp。
此外，共有74,732个单基因簇可比对至以下至少一个数据库：非冗余蛋白质序列数据库（NR）、基因本体论（GO）、京都基因与基因组百科全书（KEGG）、KEGG直系同源数据库（KO）及蛋白质家族数据库（Pfam）。本研究同时鉴定出6696个转录因子（TFs）、28,458个长链非编码RNA（lncRNAs）以及94,230组mRNA-miRNA配对关系。
中华绒螯蟹肝孢虫（Hepatospora eriocheir）是中华绒螯蟹的主要病原，可引发肝胰腺坏死病（HPND），肠道为其主要侵染靶组织。本研究旨在筛选中华绒螯蟹肠道内与中华绒螯蟹肝孢虫侵染相关的关键基因。
通过联合PacBio Iso-Seq测序与Illumina RNA测序（Illumina RNA-seq）分析，我们在感染中华绒螯蟹肝孢虫的中华绒螯蟹肠道中共鉴定出12,708个差异表达单基因簇（DEUs），其中6,696个呈上调表达，6,012个呈下调表达。
基于对上述差异表达单基因簇的生物学分析，我们识别出多个关键调控通路，包括能量代谢相关通路、细胞凋亡通路及先天免疫相关通路。随后，我们从这些差异表达单基因簇中选取12个基因，通过实时荧光定量PCR（qRT-PCR）完成了验证。本研究结果深化了我们对中华绒螯蟹转录组以及中华绒螯蟹与肝孢虫侵染之间特异性互作关系的认知。