低温胁迫下乐东拟单性木兰的转录组测序及生物信息学分析  

Transcriptome and Bioinformatics Analysis of Parakmeria lotungensis in Response to Low Temperature Stress

作　　者：汪结明 李瑞雪[1,2] 李贯虹 田敬璞 WANG Jieming;LI Ruixue;LI Guanhong;TIAN Jingpu(Hunan University of Science and Technology,Xiangtan,Hunan 411201,China;Hunan Tricholoma Agricultural Development Co.,Ltd,Xiangtan,Hunan 411201,China)

机构地区：[1]湖南科技大学,风景园林系,湖南湘潭411201 [2]湖南蘑蘑哒农业开发有限公司,湖南湘潭411201

出　　处：《复旦学报(自然科学版)》2025年第1期96-112,共17页Journal of Fudan University:Natural Science

基　　金：教育部人文社科一般项目(24YJAZH144);湖南省自然科学基金省市联合项目(2021JJ50119);湖南省社会科学成果评审委员会项目(XSP22YBZ073);湖南省企业科技特派员项目(2021GK5036)。

摘　　要：为探究乐东拟单性木兰(Parakmeria lotungensis)应对低温胁迫的响应机理,采用转录组测序技术对0℃冷胁迫下2 h、6 h、24 h和常温(25℃)对照0 h的乐东拟单性木兰的RNA文库进行转录组测序,对测序结果进行de novo拼接,筛选差异表达的unigenes,并对其进行功能注释,同时选择6个与乐东拟单性木兰抗寒相关的差异表达基因进行qRT-PCR验证。结果表明,总共获得平均长度为740.64 bp的unigene 273252个,其中注释到Uniprot、Pfam、GO、KEGG、KEGG通路、eggNOG和Nr数据库的unigenes分别有70624、25189、49622、9079、5819、6139和69070个。0℃低温胁迫24 h后差异表达的unigenes数量最多,为19231个。基于公共基因数据库BLAST比对和KEGG通路分析,发现Pfam结构域注释以富含亮氨酸重复家族(LRR)、ATPase家族和蛋白激酶家族最多,GO数据库注释发现这些差异基因显著富集在光合作用、压力应激反应、氧化应激反应、谷氨酸酶代谢、水解酶活性、脂质结合、蛋白激酶、氧化还原酶活性等条目,KEGG通路注释结果表明,与碳水化合物代谢有关的unigenes最多(1401条),显著富集在淀粉与糖代谢途经,表明这些unigenes可能参与乐东拟单性木兰应对低温胁迫的耐受性作用。随机挑选6个与乐东拟单性木兰抗寒相关的差异表达基因进行qRT-PCR验证。结果表明,这6个差异基因的qRT-PCR相对表达量(2^(-ΔΔCT))与转录组测序的相对表达量(FPKM)的动态变化趋势基本一致,进一步验证了乐东拟单性木兰转录组测序数据的重复性和可靠性。In order to reveal the adaptation mechanisms of Parakmeria lotungensis to low temperature,transcriptome sequencing analysis was performed under low temperature for 2 hours,6 hours,24 hours,and normal temperature control for 0 hours(25℃).RNA sequencing quality were evaluated,DEGs were functionally annotated and bioinformatics analyzed.Six dDEGs were selected and performed for qRT-PCR to validate the RNA-seq datas.The results showed that these clean reads were trimmed and assembled into 273252 non-redundant putative transcripts with a mean length of 590.52 bp.A total of 70624,25189,49622,9079,5819,6139 and 69070 unigenes had the most significant BLAST matches with known proteins in the Uniprot,Pfam,GO,KEGG,KEGG Pathway,eggNOG,and Nr databases respectively.The highest number of differentially expressed genes were 19231.Based on the functional annotation through BLAST with public protein database,2369 and 2173 entries of unigenes from leucine rich repeat family,ATPase family and protein kinase family were annotated in Pfam domain respectively.GO annotation showed that these unigenes were involved in biosynthetic process,cellular component and molecular function and significantly enriched in photosynthesis,response to stress,response to oxidative stress,glutamate metabolic process,hydrolase activity,lipid binding,protein kinase,oxidoreductase activity,transferase activity and cell wall organization.The KEGG path annotation results indicate that 6299 entries(41.85%)of unigenes among with them,1401 entries of unigenes were related to carbohydrate metabolism and significantly enriched in starch and sucrose metabolism pathway.This indicates that these unigenes may be involved in the tolerance of Parakmeria lotungensis to low temperature stress.Six DEGs were selected and performed for real-time fluorescence quantitative(qRT-PCR)to validate the RNA-seq data.qRT-PCR analysis results showed that the relative expression(2^(-ΔΔCT))of these six DEGs by qRT-PCR was basically consistent with the dynamic change trend of the rel

关 键 词：乐东拟单性木兰 低温胁迫 转录组 差异表达 

分 类 号：Q945.78[生物学—植物学] Q943.2