高温胁迫诱导薄荷差异表达基因的分析

Analysis of Differentially Expressed Genes in Mentha haplocalyx Briq. Induced by High Temperature Stress

目的:分析薄荷幼苗高温胁迫下的差异基因表达情况,从中挖掘薄荷幼苗的耐热功能基因,为薄荷幼苗的抗逆机制提供生物信息学依据。方法:以水培薄荷为研究对象,分别进行高温胁迫0 h(对照,CK)、3 h(40℃,HT-3 h)和6 h(40℃,HT-6 h)的3种处理,然后进行转录组测序,对比分析筛选出高温胁迫应答的基因,并对部分基因利用实时荧光定量PCR(qRT-PCR)进行验证。结果:HT-3 h vs CK和HT-6 h vs CK的共同差异表达基因(Differential expressed genes,DEGs)有1042个,其中上调基因590个,下调基因452个。对共有DEGs进行GO功能富集分析,以对p值矫正后的FDR≤0.05为阈值,得出55条GO terms功能组为显著富集,主要体现在生物过程(Biological Process,BP)方面,包括活性氧反应、对刺激的反应、热反应等;KEGG功能富集分析得出,共有611个DEGs映射到171条通路当中,主要涉及代谢途径、内质网中的蛋白质加工、核糖体、次级代谢产物的生物合成等方面。进一步挖掘响应HT-3 h和HT-6 h的共有DEGs中,上调的有33个Hsp家族的基因,主要包括19个Hsp20基因,5个Hsp90基因和7个HSPAls基因等;另有5个转录因子BES1、GRAS、HB-HD-ZIP、MYB-related、NAC亦显示上调。利用qRT-PCR对测序数据进行验证,基因的表达趋势与转录组测序结果一致,证明测序结果真实可靠。结论:薄荷通过启动活性氧代谢或氧化应激等生物进程来响应高温胁迫,并通过内质网中的蛋白质加工等途径来抵御高温伤害。上调热激蛋白Hsp20、Hsp90、HSPAls等基因和BES1、GRAS、HB-HD-ZIP、MYB-related、NAC等5个转录因子家族的相关基因,对薄荷的耐热调控具有积极意义。

Objective:The differential gene expression of peppermint seedlings under high temperature stress was analyzed to explore the heat-resistant functional genes of peppermint seedlings,so as to provide bioinformatics basis for the stress resistance mechanism of peppermint seedlings.Methods:Hydroponic peppermint was treated with high temperature stress for 0 h(control,CK),3 h(40℃,HT-3 h)and 6 h(40℃,HT-6 h),and then transcriptome sequencing was carried out.The genes responding to high temperature stress were screened by comparative analysis,and some genes were verified by real-time quantitative PCR.Results:There were 1042 differentially expressed genes(DEGs)in(HT-3 h)vs CK and(HT-6 h)vs CK,including 590 up-regulated genes and 452 down regulated genes(biological process,BP),including reactive oxygen species response,response to stimulation,thermal response,etc..KEGG function enrichment analysis shows that a total of 611 DEGs are mapped to 171 pathways,mainly involving metabolic pathways,protein processing in endoplasmic reticulum,ribosome,biosynthesis of secondary metabolites,etc.further explore the up regulation of common DEGs in response to HT-3 h and HT-6 h.There are 33 HSP family genes,mainly including 19 Hsp20 genes,5 Hsp90 genes and 7 Hspals genes.Five transcription factors BES1,GRAS,HB-HD-ZIP,MYB-related and NAC were also up-regulated.The sequencing data were verified by fluorescence quantitative PCR(qRT-PCR).The gene expression trend was consistent with the transcriptome sequencing results,which proved that the sequencing results were true and reliable.Conclusion:Peppermint can respond to high temperature stress by initiating various biological processes such as reactive oxygen species metabolism or oxidative stress,and resist high temperature injury through protein processing in endoplasmic reticulum.Up regulation of heat shock proteins Hsp20,Hsp90,HspAls and related genes of five transcription factor families such as BES1,GRAS,HB-HD-ZIP,MYB-related and NAC will be of positive significance to the heat-resistant regulation of peppermint.