短密木霉降解咪唑乙烟酸候选基因的筛选
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摘要
为探明短密木霉(Trichoderma brevicompactum)降解咪唑乙烟酸的分子机制,通过转录组学和蛋白质组学数据整合分析筛选降解咪唑乙烟酸的候选基因。结果显示:以500 mg/kg咪唑乙烟酸驯化后的短密木霉为材料,分别在基础培养基和以咪唑乙烟酸为单一碳源的培养基中培养至降解效率最高的第7天,在转录组学中差异表达的基因1 329个,其中上调703个,下调626个。用ITRAQ进行蛋白质组学分析,共鉴定到21 883条肽段和4 003个蛋白质。关联分析中,鉴定出24个在转录组和蛋白质组中均差异表达的基因,其中14个基因上调,10个基因下调。定量蛋白质和基因的表达关联系数为-0.047 2,显著差异蛋白质和显著差异基因表达关联系数为-0.142 0,蛋白质和表达变化趋势相同的基因关联系数为-0.741 3,蛋白质和表达变化趋势相反的基因关联系数为-0.791 6。关联分析鉴定的24个基因中,有2个基因具有功能注释,分别为TBU3981A(NCBInr:绿色木霉Gv29-8糖苷水解酶16家族部分mRNA)和TBU1425A(NCBInr:HEX-1),可作为降解咪唑乙烟酸的候选基因。
The candidate genes of degradating imazethapyr were screened with integration analyses of transcriptomics and proteomics data to explore the molecular mechanism of degradating imazethapyr in Trichoderma brevicompactum. The results showed that when the basal medium with 500 mg/kg mazethapyr was used as the sole carbon source to cultivate T. brevicompactum for seven days, the highest degradation efficiency was obtained. There were 1 329 genes with significant transcriptomic differences, of which 703 were up-regulated and 626 were down-regulated. 21 883 peptides and 4 003 proteins were identified by proteome analyses performed with ITRAQ. The results of association analyses showed that 24 genes with differential expression and protein abundance were correspondingly changed, of which 14 were up-regulated and 10 were down-regulated. The association coefficient between quantitative protein and gene expression was-0.047 2. The association coefficient between significant difference protein and genes with significantly differential expression was-0.142 0, with the similar trend of expression change. The association coefficient between protein and gene expression was-0.741 3.The association coefficient between protein and genes with the opposite trend of expression change was-0.791 6. Two of the 24 genes identified by association analysis had functional annotations. They are Trichoderma virens Gv29-8 glycoside hydrolase family 16 protein partial mRNA and HEX-1 gene, respectively. They can be used as candidate genes of degradating imazethapyr. It is indicated that the candidate genes of degradating imazethapyr by T. brevicompactum can be effectively screened with the association analyses of transcriptome and proteome.
The candidate genes of degradating imazethapyr were screened with integration analyses of transcriptomics and proteomics data to explore the molecular mechanism of degradating imazethapyr in Trichoderma brevicompactum. The results showed that when the basal medium with 500 mg/kg mazethapyr was used as the sole carbon source to cultivate T. brevicompactum for seven days, the highest degradation efficiency was obtained. There were 1 329 genes with significant transcriptomic differences, of which 703 were up-regulated and 626 were down-regulated. 21 883 peptides and 4 003 proteins were identified by proteome analyses performed with ITRAQ. The results of association analyses showed that 24 genes with differential expression and protein abundance were correspondingly changed, of which 14 were up-regulated and 10 were down-regulated. The association coefficient between quantitative protein and gene expression was-0.047 2. The association coefficient between significant difference protein and genes with significantly differential expression was-0.142 0, with the similar trend of expression change. The association coefficient between protein and gene expression was-0.741 3.The association coefficient between protein and genes with the opposite trend of expression change was-0.791 6. Two of the 24 genes identified by association analysis had functional annotations. They are Trichoderma virens Gv29-8 glycoside hydrolase family 16 protein partial mRNA and HEX-1 gene, respectively. They can be used as candidate genes of degradating imazethapyr. It is indicated that the candidate genes of degradating imazethapyr by T. brevicompactum can be effectively screened with the association analyses of transcriptome and proteome.
引文
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[2] 丁伟,白鹤,程茁,等.咪唑乙烟酸降解菌的分离、鉴定及其降解特性研究[J].环境科学,2008(5):1359-1362.
[3] 陈玉洁,束长龙,刘新刚,等.咪唑乙烟酸降解菌的分离、鉴定及降解特性研究[J].农药学学报,2011(4):387-393.
[4] 曹知平,许景钢,李淑芹,等.黑曲霉LZ1降解咪唑乙烟酸的特性[J].东北农业大学学报,2010(7):66-70.
[5] 吕翻洋,许泽华,毛晓洁,等.咪唑乙烟酸降解菌的分离鉴定及降解条件优化[J].生物技术通报,2017(10):163-168.
[6] 金雷,成明根,孙斌,等.芽孢杆菌QC-13对咪唑乙烟酸污染土壤的生物修复[J].江苏农业科学,2015(6):300-303.
[7] MAIER T,GUELL M,SERRANO L.Correlation of mRNA and protein in complex biological samples[J].Feebs letters,2009,583(24):3966-3973.
[8] 王青玲.一株乙羧氟草醚高效降解菌的分离鉴定、降解特性及其应用研究[D].南京:南京农业大学,2012.
[9] 靳颖华.液相色谱法测定燕窝中唾液酸含量研究[J].中国医药导刊,2011(6):1071-1072.
[10] CANOVAS A,RINCON G,ISLAS-TREJO A,et al.SNP discovery in the bovine milk transcriptome using RNA-Seq technology[J].Mammelian genome,2010,21(11/12):592-598.
[11] LI X,SUN H,PEI J,et al.De novo sequencing and comparative analysis of the blueberry transcriptome to discover putative genes related to antioxidants[J].Gene,2012,511(1):54-61.
[12] GRABHERR M G,HAAS B J,YASSOUR M,et al.Full-length transcriptome assembly from RNA-Seq data without a reference genome[J].Nature biotechnology,2011,29(7):644-652.
[13] CONESA A,G?TZ S,GARCíA-GóMEZ J M.Blast2GO:a universal tool for annotation,visualization and analysis in functional genomics research[J].Bioinformatics,2005,21(18):3674-3676.
[14] LAN P,LI W,SCHMIDT W.Complementary proteome and transcriptome profiling in phosphate-deficient arabidopsis roots reveals multiple levels of gene regulation[J].Molecular & cellular proteomics,2012,11:1156-1166.
[15] HUANG Y,MIJITI G,WANG Z,et al.Functional analysis of the class Ⅱ hydrophobin gene HFB2-6 from the biocontrol agent Trichoderma asperellum ACCC30536[J].Microbiological research,2015,171:8-20.
[16] 马进,郑钢.利用转录组测序技术鉴定紫花苜蓿根系盐胁迫应答基因[J].核农学报,2016(8):1470-1479.
[17] LACKNER D H,SCHMIDT M W,WU S,et al.Regulation of transcriptome,translation,and proteome in response to environmental stress in fission yeast[J].Genome biology,2012,13 (4):1-14.
[18] 吴松锋,朱云平,贺福初.转录组与蛋白质组比较研究进展[J].生物化学与生物物理进展,2005(2):99-105.
[19] KUBICEK C P,HERRERA-ESTRELLA A,SEIDL-SEIBOTH V,et al.Comparative genome sequence analysis underscores mycoparasitism as the ancestral life style of Trichoderma[J/OL].Genome biology,2011,12R40[2018-08-10].http://doi.org/10.1186/gb-2011-12-4-r40.
[20] 王圆,张朝晖,王蓓,等.细菌产β-1,3-1,4-葡聚糖酶的结构,功能及应用的研究进展[J].饲料工业,2005,26(2):18-20.
[21] TENNEY K,HUNT I,SWEIGARD J,et al.HEX-1,a gene unique to filamentous fungi,encodes the major protein of the Woronin body and functions as a plug for septal pores[J].Fungal genetics and biology :FG & B,2000,31(3):205-217.
[22] JEDD G,CHUA N H.A new self-assembled peroxisomal vesicle required for efficient resealing of the plasma membrane[J].Nature cell biology,2000,2(4):226-231.
[23] CURACH N C,TE’O V,GIBBS M D,et al.Isolation,characterization and expression of the HEX-1 gene from Trichoderma reesei[J].Gene,2004,331:133-140.
[24] YUAN P,JEDD G,KUMARAN D,et al.A HEX-1 crystal lattice required for Woronin body function in Neurospora crassa[J].Nature structural biology,2003,10(4):264-270.
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[27] 唐俊,陆娟,许惠芬,等.康氏木霉HEX-1蛋白的原核表达及纯化[J].西北农林科技大学学报(自然科学版),2014(10):147-151.
[28] TANG J,LIU L,HUANG X,et al.Proteomic analysis of Trichoderma atroviride mycelia stressed by organophosphate pesticide dichlorvos[J].Canadian journal of microbiology,2010,56(2):121-127.