温度胁迫对幼体大海马基因转录表达的影响

Effects of Temperature Stress on Genes’ Transcription and Expression in Juvenile Hippocampus kuda Bleeker

为探究大海马幼体在高温和低温胁迫条件下基因表达水平的变化规律,采用Illumina平台的PE150测序功能对大海马幼体肝脏进行了转录组测序。对照组、高温胁迫组和低温胁迫组测序数据组装后获得22 513个单基因簇(Unigene),N50为2 223 bp,平均长度为1 122.71 bp。与对照组相比,高温胁迫组共获得14 009个差异表达基因,其中5 543个差异表达基因上调,8 466个差异表达基因下调;低温胁迫组共获得20 030个差异基因,14 016个差异表达基因上调,6 014个差异表达基因下调。差异表达基因经KEGG数据库富集发现,高/低温胁迫均导致大海马体内抗氧化途径相关基因表达(HSP70、HSP90、SOD等)发生显著改变;另外,高温胁迫还造成大海马免疫系统相关基因(PIK3R、Akt、IL-10、TLR等)以及细胞凋亡相关基因(CYCS、CASP9、CASP3等)表达显著改变;低温胁迫造成DNA损伤修复(MSH、DDB2、XRCC2、RAD52、Ogg1、PMS2等)、脂肪酸代谢(StARD7、ApoA4、CYP51、Fadsd6等)和细胞凋亡(P21、P53、BAX等)相关基因显著上调。选择HSP70、Fadsd 6与IL-10等13个差异基因验证,RT-qPCR结果与RNA-Seq测序结果基本一致。本研究结果为深入探究大海马温度应激的调控机制奠定了一定的理论基础,而且有助于预防极端温度对大海马造成损伤。

In order to investigate the changes in gene expression levels of juvenile H. kuda under high and low temperature stress, the transcriptome sequencing of the juvenile’s liver was performed using the PE150 sequencing strategy based on the Illumina platform. Sequencing data of control group, high temperature stress group and low temperature stress group were assembled totally 22 513 single gene clusters(Unigene) were obtained, with an N50 of 2 223 bp and an average length of 1 122.71 bp. Compared with the control, 14 009 differentially-expressed genes were obtained in the high temperature group, of which 5 443 genes were up-regulated and 8 416 genes were down-regulated. Compared with the control, 20 030 differentially-expressed genes were also obtained in the low temperature group(14 016 genes were up-regulated and 6 014 genes were down-regulated). Enriched by KEGG database, the differentially-expressed genes under high/low temperature stress both caused significant changes in the antioxidant pathway-related genes(HSP70, HSP90, SOD, etc.) in juvenile H. kuda. In addition, The high temperature stress also caused significant changes in the expression of immune system-related genes(PIK3R, AKT, IL-10, TLR, etc.) and apoptosis-related genes(CYCS, CASP9, CASP3, etc.), while the low temperature stress caused significant changes of genes related to DNA damage repair(MSH, DDB2, XRCC2, RAD52, OGG1, PMS2, etc.), fatty acid metabolism(StARD7, APOA4, CYP51, Fadsd6, etc.) and cell apoptosis(P21, P53, BAX, etc.). Thirteen differentially-expressed genes such as IL-10, HSP70 and stearoyl-CoA desaturase were selected to validate the results, and the results of RT-qPCR were almost consistent with those of RNA sequencing. This work can lay a preliminary foundation for further research on the regulation mechanism of temperature stress in H. kuda, and help to prevent extreme temperature damage to the culture of H. kuda.