可可毛色二孢菌侵染后的烟草酵母双杂交cDNA文库构建
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摘要
<正>葡萄是世界上最重要的经济果树之一,据联合国粮食及农业组织统计,截止到2013年,中国葡萄栽培面积已达715.5万hm2,葡萄总产量超2 300万kg。长期以来,葡萄病害是影响葡萄产业健康发展的重要因素。近年来研究发现由葡萄座腔菌科真菌(Botryosphaeriaceae)引起的葡萄溃疡病(Botryosphaeria dieback)是葡萄上的主要枝干病
The occurrence of grapevine canker caused by pathogens of fungal family Botryosphaeriaceae is increasing in most grapevine grow ing areas w orldw ide. How ever,the mechanism of Botryosphaeriaceae-grapevine interaction is still in its infancy. In order to identify Botryosphaeriaceae resistance-related genes and targets of fungal effectors in grapevine,a yeast tw o-hybrid c DNA library of tobacco infected by Lasiodiplodia theobromae CSS-01 s w as constructed by SM ART c DNA synthesis technology. The results show ed that the titer of c DNA library w as 1.2×108 CFU·m L-1. Results of PCR amplification of 24 clones randomly selected from the c DNA library revealed that the length of inserted c DNA fragments ranged from 350 bp to 2 000 bp,w ith a recombination rate of 100%. In addition,the average insert size of the c DNA library w as 750 bp,indicating that this library is of high quality and could be applied for subsequent research.
The occurrence of grapevine canker caused by pathogens of fungal family Botryosphaeriaceae is increasing in most grapevine grow ing areas w orldw ide. How ever,the mechanism of Botryosphaeriaceae-grapevine interaction is still in its infancy. In order to identify Botryosphaeriaceae resistance-related genes and targets of fungal effectors in grapevine,a yeast tw o-hybrid c DNA library of tobacco infected by Lasiodiplodia theobromae CSS-01 s w as constructed by SM ART c DNA synthesis technology. The results show ed that the titer of c DNA library w as 1.2×108 CFU·m L-1. Results of PCR amplification of 24 clones randomly selected from the c DNA library revealed that the length of inserted c DNA fragments ranged from 350 bp to 2 000 bp,w ith a recombination rate of 100%. In addition,the average insert size of the c DNA library w as 750 bp,indicating that this library is of high quality and could be applied for subsequent research.
引文
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[14]de Jonge R,van Esse H P,Kombrink A,et al.Conserved fungal Lys M effector Ecp6 prevents chitin-triggered immunity in plants[J].Science,2010,329(5994):953-955.
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[16]Cesari S,Thilliez G,Ribot C,et al.The rice resistance protein pair RGA4/RGA5 recognizes the Magnaporthe oryzae effectors AVR-Pia and AVR1-CO39 by direct binding[J].The Plant Cell,2013,25(4):1463-1481.
[17]Liu T,Song T,Zhang X,et al.Unconventionally secreted effectors of tw o filamentous pathogens target plant salicylate biosynthesis[J].Nature Communications,2014,5:10.1038/ncomms5686.
[18]Wang X,Jiang N,Liu J,et al.The role of effectors and host immunity in plant-necrotrophic fungal interactions[J].Virulence,2014,5(7):722-732.
[2]Liu A F,Zhang X,Zhang W,et al.Establishment of a rapid virulence evaluation system in Botrysphaeria rhodina transformants(in Chinese)[J].Plant Protection(植物保护),2013,39(5):96-100.
[3]Zhang W,Yan J Y,Guo X,et al.Development of a method for protoplast preparation and regeneration of Botrysphaeria rhodina(in Chinese)[J].Plant Protection(植物保护),2012,38(5):27-30.
[4]Yan J,Xie Y,Yao S,et al.Characterization of Botryosphaeria dothidea,the causal agent of grapevine canker in China[J].Australasian Plant Pathology,2012,41(4):351-357.
[5]Yan J Y,Xie Y,Zhang W,et al.Species of Botryosphaeriaceae involved in grapevine dieback in China[J].Fungal Diversity,2013,61(1):221-236.
[6]Dissanayake A J,Zhang W,Mei L,et al.Lasiodiplodia pseudotheobromae causes pedicel and peduncle discolouration of grapes in China[J].Australasian Plant Disease Notes,2015,10(1):1-5.
[7]Dissanayake A J,Zhang W,Li X,et al.First report of Neofusicoccum mangiferae associated w ith grapevine dieback in China[J].Phytopathologia M editerranea,2015,54(2):414.
[8]Urbez-Torres J R.The status of Botryosphaeriaceae species infecting grapevines[J].Phytopathologia M editerranea,2011,50(4):5-45.
[9]Boller T,Felix G.A renaissance of elicitors:perception of microbe-associated molecular patterns and danger signals by pattern-recognition receptors[J].Annual Review of Plant Biology,2009,60:379-406.
[10]Zipfel C.Early molecular events in PAMP-triggered immunity[J].Current Opinion in Plant Biology,2009,12(4):414-420.
[11]Huang Y Y,Yang L L,Wang Y,et al.Construction and utilization of a yeast tw o-hybrid c DNA library from Arabidopsis thaliana infected by pow dery mildew(in Chinese)[J].Acta Phytopathologica Sinica(植物病理学报),2015,45(4):370-375.
[12]Wang Y Q,Li G B,Yang J,et al.Construction and application of a yeast tw o-hybrid c DNA library from rice spikelets infected w ith Ustilaginoidea virens(in Chinese)[J].Scientia Agricultura Sinica(中国农业科学),2016,49(5):865-873.
[13]Shi Y,Wang Y Z,Sun H,et al.Construction of a yeast tw o-hybrid c DNA library from melon infected w ith Cucurbit chlorotic yellows virus(in Chinese)[J].Acta Phytophylacica Sinica(植物保护学报),2014(4):509-510.
[14]de Jonge R,van Esse H P,Kombrink A,et al.Conserved fungal Lys M effector Ecp6 prevents chitin-triggered immunity in plants[J].Science,2010,329(5994):953-955.
[15]Caillaud M C,Asai S,Rallapalli G,et al.A downy mildew effector attenuates salicylic acid–triggered immunity in Arabidopsis by interacting w ith the host mediator complex[J].PLo S Biology,2013,11(12):e1001732.
[16]Cesari S,Thilliez G,Ribot C,et al.The rice resistance protein pair RGA4/RGA5 recognizes the Magnaporthe oryzae effectors AVR-Pia and AVR1-CO39 by direct binding[J].The Plant Cell,2013,25(4):1463-1481.
[17]Liu T,Song T,Zhang X,et al.Unconventionally secreted effectors of tw o filamentous pathogens target plant salicylate biosynthesis[J].Nature Communications,2014,5:10.1038/ncomms5686.
[18]Wang X,Jiang N,Liu J,et al.The role of effectors and host immunity in plant-necrotrophic fungal interactions[J].Virulence,2014,5(7):722-732.