昆虫学报 ›› 2016, Vol. 59 ›› Issue (6): 581-591.doi: 10.16380/j.kcxb.2016.06.001

• 研究论文 •    下一篇

准噶尔小胸鳖甲短时低温胁迫响应的转录组分析

库尔班·吐松, 陆雪莹, 刘小宁, 马纪*   

  1. (新疆大学生命科学与技术学院,新疆生物资源基因工程重点实验室,乌鲁木齐830046)
  • 出版日期:2016-06-20 发布日期:2016-06-20

Transcriptomic analysis of the desert beetle Microdera punctipennis (Coleoptera: Tenebrionidae) in response to short-term cold stress

 Kuerban TUSONG, LU  Xue-Ying, LIU  Xiao-Ning, MA Ji*   

  1. (Xinjiang Key Laboratory of Biological Resources and Genetic Engineering, College of Life Science and Technology, Xinjiang University, Urumqi 830046, China)
  • Online:2016-06-20 Published:2016-06-20

摘要: 【目的】拟步甲科昆虫小胸鳖甲Microdera punctipennis 是分布于中国新疆古尔班通古特沙漠的特有物种,具有很强的耐寒性,但其低温响应的分子机制尚不明确。本研究旨在利用转录组测序技术丰富小胸鳖甲的已知基因信息,分析在短时低温胁迫下,上调表达基因的主要类群及参与的主要代谢通路。【方法】利用Trinity软件对分别在4℃低温和25℃(对照)下处理3 h的小胸鳖甲成虫RNA-Seq数据进行从头组装,利用Blast2go软件进行基因注释。通过DESeq和GFOLD软件分析差异表达基因,并对上调表达基因进行GO(Gene Ontology)和KEGG(Kyoto Encyclopedia of Genes and Genomes)代谢途径富集分析。【结果】测序过滤后得到68 165 001个有效读序,51 712个平均长度为984 bp的非冗余基因,其中29 925个基因(约57.87%)有同源序列(E-value<10-5),与拟步甲科的赤拟谷盗Tribolium castaneum相应基因相似性最高(核苷酸序列一致性为72.75%)。低温响应上调表达基因有514个,富集到35个GO类群,18个KEGG途径。其中胁迫响应类群、生物调节类群和免疫系统过程类群都占有重要比例,嘌呤代谢、硫胺素代谢、糖酵解/糖异生等代谢通路显著富集。在胁迫响应类群中,热激蛋白Hsp90基因、超氧化物歧化酶(SOD)基因和钙联接蛋白Calnexin基因等8个非生物胁迫相关基因显著上调表达。【结论】荒漠昆虫小胸鳖甲低温转录组揭示,在4℃冷驯化过程中,代谢过程、胁迫响应、生物调节和免疫系统等生物学过程相关基因表达显著上调,其中几个非生物胁迫响应基因提示小胸鳖甲可能从分子伴侣、细胞周期阻滞、线粒体稳定性、抗氧化胁迫等多方面应对低温胁迫。KEGG富集分析所揭示的小胸鳖甲在低温下的转录组代谢通路与其他物种有较大差异。研究结果为后续生物信息学分析及小胸鳖甲耐寒关键基因的发掘及耐寒机制的揭示提供了基础数据。

关键词: 小胸鳖甲, 低温胁迫, 冷驯化, 转录组, 差异基因

Abstract: 【Aim】 Tenebrionid insect Microdera punctipennis is an endemic species in Gurbantünggüt Desert, Xinjiang, China. It has very strong cold hardiness, but the molecular mechanisms by which M. punctipennis responses to low temperature are still unclear. This study aims to enrich the genetic information of M. punctipennis , and to analyze the functional groups of the most up-regulated genes under short-term cold stress and the pathways which they may involve in. 【Methods】 Transcriptomic sequencing reads from M. punctipennis adults exposed to 4℃ as low temperature treatment and 25℃ as the control, respectively, for 3 h were de novo assembled by using Trinity software. Gene annotation was conducted by using Blast2go software. The differentially expressed genes were screened by using DESeq and GFOLD softwares. For the up-regulated genes, GO term enrichment and KEGG metabolic pathway analysis were performed using Blast2go software. 【Results】 In total, 51 712 non-redundant transcripts with the average length of 984 bp were assembled, of which the expression levels of 514 genes were up-regulated and 91 genes down-regulated in response to cold stress. The up-regulated ones were enriched to 35 GO terms and 18 KEGG pathways, showing that stress-related GO terms, biological regulation and immune system are enriched significantly. Pathways such as purine metabolism, thiamine metabolism, and glycolysis/gluconeogenesis are significantly enriched. In the stress responsive GO term, the expression levels of eight abiotic related genes, including heat shock protein (Hsp90) gene, superoxide dismutase (SOD) gene, calnexin gene, etc., were up-regulated significantly. 【Conclusion】 The transcriptomic data of the desert beetle M. punctipennis reveal that the expressions of genes related to the biological processes like metabolic process, stress response, biological regulation and immune system are significantly up-regulated during cold acclimation at 4℃. The metabolic pathways revealed by KEGG enrichment show that low temperature transcriptome of M. punctipennis is much different from those of other species. Our results provide basic data for successive bioinformatics analysis and mining the key genes in cold tolerance for M. punctipennis as well as for elucidation of the related molecular mechanisms.

Key words: Microdera punctipennis, low temperature stress, cold acclimation, transcriptome, differentially expressed genes