双孢蘑菇6个菌株不同发育阶段的转录组分析
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摘要
【目的】通过多个双孢蘑菇菌株不同发育阶段的差异转录组分析,为进一步验证双孢蘑菇发育相关基因及探讨其分子机理奠定基础。【方法】对双孢蘑菇主栽品种As2796及其亲本02、8213,其回交子代W192,以及国外野生菌株ARP159、国内野生菌株AgLH830共6个具有重要代表性的菌株子实体原基期、幼菇期、采摘期、开伞期等4个不同发育阶段共24个样品进行转录组测序,并与双孢蘑菇参考基因组序列进行比对,根据比对结果进行各基因在不同样品中的表达量分析及差异表达基因识别,发掘新基因与共同基因的差异表达,并进行各数据库的基因功能注释。【结果】结果共鉴定到10 660个转录本,发掘新基因677个,其中237个得到功能注释。与原基期相比,6个菌株在幼菇期、采摘期和开伞期分别有49、82、73个共同差异表达基因,其中有13个是相同的基因。【结论】发现了一批在双孢蘑菇子实体不同发育阶段具有显著差异表达的基因,筛选出不同菌株不同阶段的共有差异基因,对双孢蘑菇子实体发育中重要的差异基因进行了注释与探讨。
【Objective】Transcriptome analysis was conducted on Agaricus bisporus at 4 developmental stages to identify the genes associated with and decipher the molecular mechanism involving the fungal development. 【Method】From 6 representative strains of A. bisporus including main cultivar As2796, its parents 02 and 8213, its backcrossing offspring W192, foreign wild strain ARP159, and domestic wild strain AgLH830, at primordium, young, harvesting, and opening stages, 24 fruiting body specimens were collected for transcriptome sequence analysis. By aligning them against the reference genome sequence of A. bisporus, the genes that were differentially expressed were identified. Both unique and common differentially expressed genes(DEGs) were clearly exposed to be annotated using the databases to determine their specific functions. 【Result】 Among the 10 660 transcripts obtained, 677 genes were unique with 237 functionally annotated. The tested A. bisporus shared 49 common DEGs between primordium and young stages, 82 between primordium and harvest stages, and 73 between primordium and pileus opening stages. And, 13 genes were found commonly present in the various strains. 【Conclusion】 Both unique and common DEGs in A. bisporus at the 4 developmental stages were identified and annotated in the study.
【Objective】Transcriptome analysis was conducted on Agaricus bisporus at 4 developmental stages to identify the genes associated with and decipher the molecular mechanism involving the fungal development. 【Method】From 6 representative strains of A. bisporus including main cultivar As2796, its parents 02 and 8213, its backcrossing offspring W192, foreign wild strain ARP159, and domestic wild strain AgLH830, at primordium, young, harvesting, and opening stages, 24 fruiting body specimens were collected for transcriptome sequence analysis. By aligning them against the reference genome sequence of A. bisporus, the genes that were differentially expressed were identified. Both unique and common differentially expressed genes(DEGs) were clearly exposed to be annotated using the databases to determine their specific functions. 【Result】 Among the 10 660 transcripts obtained, 677 genes were unique with 237 functionally annotated. The tested A. bisporus shared 49 common DEGs between primordium and young stages, 82 between primordium and harvest stages, and 73 between primordium and pileus opening stages. And, 13 genes were found commonly present in the various strains. 【Conclusion】 Both unique and common DEGs in A. bisporus at the 4 developmental stages were identified and annotated in the study.
引文
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[20]HANSEN K D,WU Z J,IRIZARRY R A,et al.Sequencing technology does not eliminate biological variability[J].Nature Biotechnology,2011,29(7):572-573.
[21]吴小梅,张昕,李南羿.双孢蘑菇子实体不同发育时期的转录组分析[J].菌物学报,2017,36(2):193-203.WU X M,ZHANG X,LI N Y.Transcriptome analysis of Agaricus bisporus fruiting at different stages[J].Mycosystema,2017,36(2):193-203.(in Chinese)
[2]TEICHERT I,WOLFF G,KüCK U,et al.Combining laser microdissection and RNA-seq to chart the transcriptional landscape of fungal development[J].BMC Genomics,2012,13(1):511-529.
[3]杨芳,许波,李俊俊,等.鸡枞菌转录组分析揭示其对木质纤维素的降解功能[J].微生物学报,2012,52(4):466-477.YANG F,XU B,LI J J,et al.Transcriptome analysis of Termitomyces albuminosus reveals the biodegradation of lignocellulose[J].Acta Microbiologica Sinica,2012,52(4):466-477.(in Chinese)
[4]CHEN L F,GONG Y H,CAI Y L,et al.Genome sequence of the edible cultivated mushroom Lentinula edodes (Shiitake) reveals insights into lignocellulose degradation[J].PloS One,2016,11(8),e0160336.
[5]FU Y P,DAI Y T,YANG C T,et al.Comparative transcriptome analysis identified candidate genes related to Bailinggu mushroom formation and genetic markers for genetic analyses and breeding[J].Scientific Reports,2017,7(1):9266.
[6]陈美元.双孢蘑菇子实体原基与菇蕾蛋白质表达变化分析[J],食用菌学报,2012,19(3):15-20.CHEN M Y.Differential Expression of Proteins During the Primordium and Button Stages of Agaricus bisporus[J].Acta Edulis Fungi,2012,19(3):15-20.(in Chinese)
[7]陈美元,廖剑华,李洪荣,等.双孢蘑菇子实体发育后期差异表达蛋白质分析[J],菌物学报,2013,32(5):855-861.CHEN M Y,LIAO J H,LI H R,et al.Analysis of differentially expressed proteins in later developing stage fruitbody of Agaricus bisporus[J].Mycosystema,2013,32(5):855-861.(in Chinese)
[8]陈美元,廖剑华,李洪荣,等.双孢蘑菇子实体发育差异蛋白质组分析[J],菌物学报,2015,34(6):1153-1164.CHEN M Y,LIAO J H,LI H R,et al.Developmental proteomics analysis of the button mushroom Agaricus bisporus[J].Mycosystema,2015,34(6):1153-1164.(in Chinese)
[9]CHEN M Y,LIAO J H,LI H R,et al.iTRAQ-MS/MS proteomic analysis reveals differentially expressed proteins during post-harvest maturation of the white button mushroom Agaricus bisporus[J].Current Microbiology,2017,74(5):641-649.
[10]施肖堃,蔡志欣,郭仲杰,等.双孢蘑菇As2796子实体发育转录组测序分析[J],福建农业学报,2018,33 (3):282-287.SHI X K,CAI Z X,GUO Z J,et al.Analysis of Agaricus bisporus Fruitbody Development by Transcriptome Sequencing[J].Fujian Journal of Agricultural Sciences,2018,33 (3):282-287.(in Chinese)
[11]蔡丹凤,蔡志欣,陈美元,等.茯苓菌落褐变的转录组测序分析[J],广州中医药大学学报,2017,34(2):245-249.CAI D F,CAI Z X,CHEN M Y,et al.Analysis of Poria cocos Mycelia Browning by Transcriptome Sequencing[J].Journal of Guangzhou University of Traditional Chinese Medicine,2017,34(2):245-249.(in Chinese)
[12]MORTAZAVI A,WILLIAMS B A,McCUE K,et al.Mapping and quantifying mammalian transcriptomes by RNA-Seq[J].Nature Methods,2008,5(7):621-628.
[13]KIM D,PERTEA G,TRAPNELL C,et al.TopHat2:accurate alignment of transcriptomes in the presence of insertions,deletions and gene fusions[J].Genome Biology,2013,14:R36.
[14]ALTSCHUL S F,MADDEN T L,ZHANG J,et al.Gapped BLAST and PSI BLAST:A New Generation of Protein Database Search Programs[J].Nucleic Acids Research,1997,25(17):3389-3402.
[15]TATUSOV R L,GALPERIN M Y,NATALE D A.The COG database:a tool for genome scale analysis of protein functions and evolution[J].Nucleic Acids Research,2000,28(1):33-36.
[16]ASHBURNER M,BALL C A,BLAKE J A,et al.Gene ontology:tool for the unification of biology[J].Nature Genetics,2000,25(1):25-29.
[17]KOONIN E V,FEDOROVA N D,JACKSON J D,et al.A comprehensive evolutionary classification of proteins encoded in complete eukaryotic genomes[J].Genome Biology,2004,5(2):77.
[18]APWEILER R,BAIROCH A,WU C H,et al.UniProt:the universal protein knowledgebase[J].Nucleic acids research,2004,32:115-119.
[19]DENG Y Y,LI J Q,WU S F,et al.Integrated nr Database in Protein Annotation System and Its Localization[J].Computer Engineering,2006,32(5):71-74.
[20]HANSEN K D,WU Z J,IRIZARRY R A,et al.Sequencing technology does not eliminate biological variability[J].Nature Biotechnology,2011,29(7):572-573.
[21]吴小梅,张昕,李南羿.双孢蘑菇子实体不同发育时期的转录组分析[J].菌物学报,2017,36(2):193-203.WU X M,ZHANG X,LI N Y.Transcriptome analysis of Agaricus bisporus fruiting at different stages[J].Mycosystema,2017,36(2):193-203.(in Chinese)
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