Transcriptomic analysis of the phytopathogenic oomycete Phytophthora cactorum provides insights into infection-related effectors

BMC Genomics

Jing-You Xu¹  Yun-Hui Tong¹  Yan-Peng Li¹  Qi-Yuan Li¹  Yu-Ping Xing¹  Bo-Yue Zhang¹  Xiao-Ren Chen¹ 

[1] College of Horticulture and Plant Protection, Yangzhou University, Yangzhou 225009, China

关键词: Polymorphism;    Plant cell death;    Transient expression;    Transcriptional profile;    WY-domain;    Effector;    TribeMCL;    Transcriptome;    Phytophthora cactorum;   

Others  :  1092431
DOI  :  10.1186/1471-2164-15-980

 received in 2014-03-31, accepted in 2014-10-29,  发布年份 2014

【 摘 要 】

Background

Phytophthora cactorum, a hemibiotrophic oomycete pathogen, can cause destructive diseases on numerous crops worldwide, leading to essential economic losses every year. However, little has been known about its molecular pathogenicity mechanisms. To gain insight into its repertoire of effectors, the P. cactorum transcriptome was investigated using Illumina RNA-seq.

Results

We first demonstrated an in vitro inoculation method that can be used to mimic natural cyst germination on host plants. Over 28 million cDNA reads were obtained for five life cycle stages (mycelium, sporangium, zoospore, cyst and germinating cyst) and de novo assembled into 21,662 unique genes. By comparisons with 11 public databases, 88.99% of the unique genes were annotated, including 15,845 mapped to the gene models of the annotated relative Phytophthora infestans. Using TribeMCL, 5,538 gene families conserved across P. cactorum and other three completely sequenced Phytophthora pathogen species were determined. In silico analyses revealed that 620 P. cactorum effector homologues including 94 RXLR effector candidates matched known or putative virulence genes in other oomycetes. About half of the RXLR effector candidates were predicted to share a conserved structure unit, termed the WY-domain fold. A subset of the effector genes were checked and validated by PCR amplification. Transcriptional experiments indicated that effector genes were differentially expressed during the life cycle and host infection stages of P. cactorum. Ectopic expression in Nicotiana benthamiana revealed that RXLR, elicitin and NLP effectors can trigger plant cell death. These effectors are highly conserved across oomycete species. Single nucleotide polymorphisms for RXLR effectors were detected in a collection of P. cactorum isolates from different countries and hosts.

Conclusions

This study demonstrates the comprehensive sequencing, de novo assembly, and analyses of the transcriptome of P. cactorum life cycle stages. In the absence of genome sequence, transcriptome data is important for infection-related gene discovery in P. cactorum, as demonstrated here for the effector genes. The first look at the transcriptome and effector arsenal of P. cactorum provides valuable data to elucidate the pathogenicity basis of this broad-host-range pathogen.

【 授权许可】

   
2014 Chen et al.; licensee BioMed Central Ltd.

【 图 表 】

Figure 1.

Figure 2.

Figure 3.

Figure 4.

Figure 5.

Figure 6.

Figure 7.

【 参考文献 】

• [1]Erwin DC, Ribeiro OK: Phytophthora Diseases Worldwide. St. Paul: APS Press; 1996.
• [2]Benson DM, Jones RK: Diseases of Woody Ornamentals and Trees in Nurseries. St. Paul: APS Press; 2001.
• [3]Hardham AR: Cell biology of plant–oomycete interactions. Cell Microbiol 2007, 9:31-39.
• [4]Shan W, Marshall JS, Hardham AR: Gene expression in germinated cysts of Phytophthora nicotianae. Mol Plant Pathol 2004, 5:317-330.
• [5]Torto-Alalibo TA, Tripathy S, Smith BM, Arredondo FD, Zhou L, Li H, Chibucos MC, Qutob D, Gijzen M, Mao C, Sobral BW, Waugh ME, Mitchell TK, Dean RA, Tyler BM: Expressed sequence tags from Phytophthora sojae reveal genes specific to development and infection. Mol Plant-Microbe Interact 2007, 20:781-793.
• [6]Judelson HS, Ah-Fong AM, Aux G, Avrova AO, Bruce C, Cakir C, da Cunha L, Grenville-Briggs L, Latijnhouwers M, Ligterink W, Meijer HJ, Roberts S, Thurber CS, Whisson SC, Birch PR, Govers F, Kamoun S, van West P, Windass J: Gene expression profiling during asexual development of the late blight pathogen Phytophthora infestans reveals a highly dynamic transcriptome. Mol Plant-Microbe Interact 2008, 21:433-447.
• [7]Savidor A, Donahoo RS, Hurtado-Gonzales O, Land ML, Shah MB, Lamour KH, McDonald WH: Cross-species global proteomics reveals conserved and unique processes in Phytophthora sojae and Phytophthora ramorum. Mol Cell Proteomics 2008, 7:1501-1516.
• [8]Chen X, Klemsdal SS, Brurberg MB: Identification and analysis of Phytophthora cactorum genes up-regulated during cyst germination and strawberry infection. Curr Genet 2011, 57:297-315.
• [9]Chen XR, Xing YP, Li YP, Tong YH, Xu JY: RNA-Seq reveals infection-related gene expression changes in Phytophthora capsici. PLoS One 2013, 8:e74588.
• [10]Kamoun S: A catalogue of the effector secretome of plant pathogenic oomycetes. Annu Rev Phytopathol 2006, 44:41-60.
• [11]Wawra S, Belmonte R, Löbach L, Saraiva M, Willems A, van West P: Secretion, delivery and function of oomycete effector proteins. Curr Opin Microbiol 2012, 15:685-691.
• [12]Tyler BM, Tripathy S, Zhang X, Dehal P, Jiang RH, Aerts A, Arredondo FD, Baxter L, Bensasson D, Beynon JL, Chapman J, Damasceno CM, Dorrance AE, Dou D, Dickerman AW, Dubchak IL, Garbelotto M, Gijzen M, Gordon SG, Govers F, Grunwald NJ, Huang W, Ivors KL, Jones RW, Kamoun S, Krampis K, Lamour KH, Lee MK, McDonald WH, Medina M, et al.: Phytophthora genome sequences uncover evolutionary origins and mechanisms of pathogenesis. Science 2006, 313:1261-1266.
• [13]Haas BJ, Kamoun S, Zody MC, Jiang RH, Handsaker RE, Cano LM, Grabherr M, Kodira CD, Raffaele S, Torto-Alalibo T, Bozkurt TO, Ah-Fong AM, Alvarado L, Anderson VL, Armstrong MR, Avrova A, Baxter L, Beynon J, Boevink PC, Bollmann SR, Bos JI, Bulone V, Cai G, Cakir C, Carrington JC, Chawner M, Conti L, Costanzo S, Ewan R, Fahlgren N, et al.: Genome sequence and analysis of the Irish potato famine pathogen Phytophthora infestans. Nature 2009, 461:393-398.
• [14]Lamour KH, Mudge J, Gobena D, Hurtado-Gonzales OP, Schmutz J, Kuo A, Miller NA, Rice BJ, Raffaele S, Cano LM, Bharti AK, Donahoo RS, Finley S, Huitema E, Hulvey J, Platt D, Salamov A, Savidor A, Sharma R, Stam R, Storey D, Thines M, Win J, Haas BJ, Dinwiddie DL, Jenkins J, Knight JR, Affourtit JP, Han CS, Chertkov O, et al.: Genome sequencing and mapping reveal loss of heterozygosity as a mechanism for rapid adaptation in the vegetable pathogen Phytophthora capsici. Mol Plant-Microbe Interact 2012, 25:1350-1360.
• [15]Kunjeti SG, Evans TA, Marsh AG, Gregory NF, Kunjeti S, Meyers BC, Kalavacharla VS, Donofrio NM: RNA-Seq reveals infection-related global gene changes in Phytophthora phaseoli, the causal agent of lima bean downy mildew. Mol Plant Pathol 2012, 13:454-466.
• [16]Ye W, Wang X, Tao K, Lu Y, Dai T, Dong S, Dou D, Gijzen M, Wang Y: Digital gene expression profiling of the Phytophthora sojae transcriptome. Mol Plant Microbe Interact 2011, 24:1530-1539.
• [17]Jupe J, Stam R, Howden AJ, Morris JA, Zhang R, Hedley PE, Huitema E: Phytophthora capsici-tomato interaction features dramatic shifts in gene expression associated with a hemi-biotrophic lifestyle. Genome Biol 2013, 14:R63. BioMed Central Full Text
• [18]Stassen JH, Van den Ackerveken G: How do oomycete effectors interfere with plant life? Curr Opin Plant Biol 2011, 14:407-414.
• [19]Gijzen M, Nurnberger T: Nep1-like proteins from plant pathogens: recruitment and diversification of the NPP1 domain across taxa. Phytochem 2006, 67:1800-1807.
• [20]Ottmann C, Luberacki B, Kufner I, Koch W, Brunner F, Weyand M, Mattinen L, Pirhonen M, Anderluh G, Seitz HU, Nürnberger T, Oecking C: A common toxin fold mediates microbial attack and plant defense. Proc Natl Acad Sci U S A 2009, 106:10359-10364.
• [21]Orsomando G, Brunetti L, Pucci K, Ruggeri B, Ruggieri S: Comparative structural and functional characterization of putative protein effectors belonging to the PcF toxin family from Phytophthora spp. Protein Sci 2011, 20:2047-2059.
• [22]Orsomando G, Lorenzi M, Raffaelli N, Dalla Rizza M, Mezzetti B, Ruggieri S: Phytotoxic protein PcF, purification, characterization, and cDNA sequencing of a novel hydroxyproline-containing factor secreted by the strawberry pathogen Phytophthora cactorum. J Biol Chem 2001, 276:21578-21584.
• [23]Rehmany AP, Gordon A, Rose LE, Allen RL, Armstrong MR, Whisson SC, Kamoun S, Tyler BM, Birch PR, Beynon JL: Differential recognition of highly divergent downy mildew avirulence gene alleles by RPP1 resistance genes from two Arabidopsis lines. Plant Cell 2005, 17:1839-1850.
• [24]Birch PR, Rehmany AP, Pritchard L, Kamoun S, Beynon JL: Trafficking arms: oomycete effectors enter host plant cells. Trends Microbiol 2006, 14:8-11.
• [25]Dou D, Kale SD, Wang X, Jiang RH, Bruce NA, Arredondo FD, Zhang X, Tyler BM: RXLR-mediated entry of Phytophthora sojae effector Avr1b into soybean cells does not require pathogen-encoded machinery. Plant Cell 2008, 20:1930-1947.
• [26]Kale SD, Gu B, Capelluto DG, Dou D, Feldman E, Rumore A, Arredondo FD, Hanlon R, Fudal I, Rouxel T, Lawrence CB, Shan W, Tyler BM: External lipid PI3P mediates entry of eukaryotic pathogen effectors into plant and animal host cells. Cell 2010, 142:284-295.
• [27]Kelley BS, Lee S, Damasceno CMB, Chakravarthy S, Kim BD, Martin GB, Rose JK: A secreted effector protein (SNE1) from Phytophthora infestans is a broadly acting suppressor of programmed cell death. Plant J 2010, 62:357-366.
• [28]Bailey K, Cevik V, Holton NJ, Byrne-Richardson J, Sohn KH, Coates M, Woods-Tör A, Aksoy HM, Hughes L, Baxter L, Jones JD, Beynon J, Holub EB, Tör M: Molecular cloning of ATR5(Emoy2) from Hyaloperonospora arabidopsidis, an avirulence determinant that triggers RPP5-mediated defense in Arabidopsis. Mol Plant-Microbe Interact 2011, 24:827-838.
• [29]Tian M, Win J, Savory E, Burkhardt A, Held M, Brandizzi F, Day B: 454 genome sequencing of Pseudoperonospora cubensis reveals effector proteins with a putative QXLR translocation motif. Mol Plant-Microbe Interact 2011, 24:543-553.
• [30]Petre B, Kamoun S: How do filamentous pathogens deliver effector proteins into plant cells? PLoS Biol 2014, 12:e1001801.
• [31]Dong S, Qutob D, Tedman-Jones J, Kuflu K, Wang Y, Tyler BM, Gijzen M: The Phytophthora sojae avirulence locus Avr3c encodes a multi-copy RXLR effector with sequence polymorphisms among pathogen strains. PLoS One 2009, 4:e5556.
• [32]Dong S, Yin W, Kong G, Yang X, Qutob D, Chen Q, Kale SD, Sui Y, Zhang Z, Dou D, Zheng X, Gijzen M, Tyler BM, Wang Y: Phytophthora sojae avirulence effector Avr3b is a secreted NADH and ADP-ribose pyrophosphorylase that modulates plant immunity. PLoS Pathog 2011, 7:e1002353.
• [33]Na R, Yu D, Qutob D, Zhao J, Gijzen M: Deletion of the Phytophthora sojae avirulence gene Avr1d causes gain of virulence on Rps1d. Mol Plant-Microbe Interact 2013, 26:969-976.
• [34]Yin W, Dong S, Zhai L, Lin Y, Zheng X, Wang Y: The Phytophthora sojae Avr1d gene encodes an RxLR-dEER effector with presence and absence polymorphisms among pathogen strains. Mol Plant-Microbe Interact 2013, 26:958-968.
• [35]Jiang RHY, Tripathy S, Govers F, Tyler BM: RXLR effector reservoir in two Phytophthora species is dominated by a single rapidly evolving superfamily with more than 700 members. Proc Natl Acad Sci U S A 2008, 105:4874-4879.
• [36]Qutob D, Tedman-Jones J, Dong S, Kuflu K, Pham H, Wang Y, Dou D, Kale SD, Arredondo FD, Tyler BM, Gijzen M: Copy number variation and transcriptional polymorphisms of Phytophthora sojae RXLR effector genes Avr1a and Avr3a. PLoS One 2009, 4:e5066.
• [37]Schornack S, Huitema E, Cano LM, Bozkurt TO, Oliva R, Van Damme M, Schwizer S, Raffaele S, Chaparro-Garcia A, Farrer R, Segretin ME, Bos J, Haas BJ, Zody MC, Nusbaum C, Win J, Thines M, Kamoun S: Ten things to know about oomycete effectors. Mol Plant Pathol 2009, 10:795-803.
• [38]Boutemy LS, King SR, Win J, Hughes RK, Clarke TA, Blumenschein TM, Kamoun S, Banfield MJ: Structures of Phytophthora RXLR effector proteins: a conserved but adaptable fold underpins functional diversity. J Biol Chem 2011, 286:35834-35842.
• [39]Win J, Krasileva KV, Kamoun S, Shirasu K, Staskawicz BJ, Banfield MJ: Sequence divergent RXLR effectors share a structural fold conserved across plant pathogenic oomycete species. PLoS Pathog 2012, 8:e1002400.
• [40]Torto TA, Li S, Styer A, Huitema E, Testa A, Gow NA, van West P, Kamoun S: EST mining and functional expression assays identify extracellular effector proteins from the plant pathogen Phytophthora. Genome Res 2003, 13:1675-1685.
• [41]Schornack S, van Damme M, Bozkurt TO, Cano LM, Smoker M, Thines M, Gaulin E, Kamoun S, Huitema E: Ancient class of translocated oomycete effectors targets the host nucleus. Proc Natl Acad Sci U S A 2010, 107:17421-17426.
• [42]Stam R, Howden AJM, Delgado-Cerezo M, Amaro TM MM, Motion GB, Pham J, Huitema E: Characterization of cell death inducing Phytophthora capsici CRN effectors suggests diverse activities in the host nucleus. Front Plant Sci 2013, 4:387.
• [43]Liu T, Ye W, Ru Y, Yang X, Gu B, Tao K, Lu S, Dong S, Zheng X, Shan W, Wang Y, Dou D: Two host cytoplasmic effectors are required for pathogenesis of Phytophthora sojae by suppression of host defenses. Plant Physiol 2011, 155:490-501.
• [44]van Damme M, Bozkurt TO, Cakir C, Schornack S, Sklenar J, Jones AM, Kamoun S: The Irish potato famine pathogen Phytophthora infestans translocates the CRN8 kinase into host plant cells. PLoS Pathog 2012, 8:e1002875.
• [45]Resjö S, Ali A, Meijer HJ, Seidl MF, Snel B, Sandin M, Levander F, Govers F, Andreasson E: Quantitative label-free phosphoproteomics of six different life stages of the late blight pathogen Phytophthora infestans reveals abundant phosphorylation of members of the CRN effector family. J Proteome Res 2014, 13:1848-1859.
• [46]Huet JC, Mansion M, Pernollet JC: Amino acid sequence of the α-elicitin secreted by Phytophthora cactorum. Phytochemistry 1993, 34:1261-1264.
• [47]Orsomando G, Lorenzi M, Ferrari E, de Chiara C, Spisni A, Ruggieri S: PcF protein from Phytophthora cactorum and its recombinant homologous elicit phenylalanine ammonia lyase activation in tomato. Cell Mol Life Sci 2003, 60:1470-1476.
• [48]Nicastro G, Orsomando G, Ferrari E, Manconi L, Desario F, Amici A, Naso A, Carpaneto A, Pertinhez TA, Ruggieri S, Spisni A: Solution structure of the phytotoxic protein PcF: the first characterized member of the Phytophthora PcF toxin family. Protein Sci 2009, 18:1786-1791.
• [49]As-sadi F, Carrere S, Gascuel Q, Hourlier T, Rengel D, Le Paslier MC, Bordat A, Boniface MC, Brunel D, Gouzy J, Godiard L, Vincourt P: Transcriptomic analysis of the interaction between Helianthus annuus and its obligate parasite Plasmopara halstedii shows single nucleotide polymorphisms in CRN sequences. BMC Genomics 2011, 12:498. BioMed Central Full Text
• [50]Kemen E, Gardiner A, Schultz-Larsen T, Kemen AC, Balmuth AL, Robert-Seilaniantz A, Bailey K, Holub E, Studholme DJ, Maclean D, Jones JD: Gene gain and loss during evolution of obligate parasitism in the white rust pathogen of Arabidopsis thaliana. PLoS Biol 2011, 9:e1001094.
• [51]Savory EA, Adhikari BN, Hamilton JP, Vaillancourt B, Buell CR, Day B: mRNA-Seq analysis of the Pseudoperonospora cubensis transcriptome during cucumber (Cucumis sativus L.) infection. PLoS One 2012, 7:e35796.
• [52]Stassen JH, Seidl MF, Vergeer PW, Nijman IJ, Snel B, Cuppen E, Van den Ackerveken G: Effector identification in the lettuce downy mildew Bremia lactucae by massively parallel transcriptome sequencing. Mol Plant Pathol 2012, 13:719-731.
• [53]Araki F, Miyagi Y: Effects of fungicides on penetration by Pyricularia oryzae as evaluated by an improved cellophane method. J Pestic Sci 1977, 2:457-461.
• [54]Mortazavi A, Williams BA, McCue K, Schaeffer L, Wold B: Mapping and quantifying mammalian transcriptomes by RNA-Seq. Nat Methods 2008, 5:621-628.
• [55]Parra G, Bradnam K, Korf I: CEGMA: a pipeline to accurately annotate core genes in eukaryotic genomes. Bioinformatics 2007, 23:1061-1067.
• [56]Win J, Morgan W, Bos J, Krasileva KV, Cano LM, Chaparro-Garcia A, Ammar R, Staskawicz BJ, Kamoun S: Adaptive evolution has targeted the C-terminal domain of the RXLR effectors of plant pathogenic oomycetes. Plant Cell 2007, 19:2349-2369.
• [57]Blair JE, Coffey MD, Park S-Y, Geiser DM, Kang S: A multi-locus phylogeny for Phytophthora utilizing markers derived from complete genome sequences. Fungal Genet Biol 2008, 45:266-277.
• [58]Stam R, Jupe J, Howden AJ, Morris JA, Boevink PC, Hedley PE, Huitema E: Identification and characterisation CRN effectors in Phytophthora capsici shows modularity and functional diversity. PLoS One 2013, 8:e59517.
• [59]Bos JIB, Kanneganti TD, Young C, Cakir C, Huitema E, Win J, Armstrong MR, Birch PR, Kamoun S: The C-terminal half of Phytophthora infestans RXLR effector AVR3a is sufficient to trigger R3a-mediated hypersensitivity and suppress INF1-induced cell death in Nicotiana benthamiana. Plant J 2006, 48:165-176.
• [60]Dou D, Kale SD, Wang X, Chen Y, Wang Q, Wang X, Jiang RH, Arredondo FD, Anderson RG, Thakur PB, McDowell JM, Wang Y, Tyler BM: Conserved C-terminal motifs required for avirulence and suppression of cell death by Phytophthora sojae effector Avr1b. Plant Cell 2008, 20:1118-1133.
• [61]Oh SK, Young C, Lee M, Oliva R, Bozkurt TO, Cano LM, Win J, Bos JI, Liu HY, van Damme M, Morgan W, Choi D, Van der Vossen EA, Vleeshouwers VG, Kamoun S: In planta expression screens of Phytophthora infestans RXLR effectors reveal diverse phenotypes, including activation of the Solanum bulbocastanum disease resistance protein Rpi-blb2. Plant Cell 2009, 21:2928-2947.
• [62]Wang Q, Han C, Ferreira AO, Yu X, Ye W, Tripathy S, Kale SD, Gu B, Sheng Y, Sui Y, Wang X, Zhang Z, Cheng B, Dong S, Shan W, Zheng X, Dou D, Tyler BM, Wang Y: Transcriptional programming and functional interactions within the Phytophthora sojae RXLR effector repertoire. Plant Cell 2011, 23:2064-2086.
• [63]Kamoun S, van West P, de Jong AJ, de Groot KE, Vleeshouwers VG, Govers F: A gene encoding a protein elicitor of Phytophthora infestans is down-regulated during infection of potato. Mol Plant Microbe Interact 1997, 10:13-20.
• [64]Yang Z: PAML 4: phylogenetic analysis by maximum likelihood. Mol Biol Evol 2007, 24:1586-1591.
• [65]Ebbole DJ: Magnaporthe as a model for understanding host-pathogen interactions. Annu Rev Phytopathol 2007, 45:437-456.
• [66]Quirino BF, Bent AF: Deciphering host resistance and pathogen virulence: The Arabidopsis/Pseudomonas interaction as a model. Mol Plant Pathol 2003, 4:517-530.
• [67]Davis TM, Yu H: A linkage map of the diploid strawberry Fragaria vesca. J Hered 1997, 88:215-221.
• [68]Shulaev V, Sargent DJ, Crowhurst RN, Mockler TC, Folkerts O, Delcher AL, Jaiswal P, Mockaitis K, Liston A, Mane SP, Burns P, Davis TM, Slovin JP, Bassil N, Hellens RP, Evans C, Harkins T, Kodira C, Desany B, Crasta OR, Jensen RV, Allan AC, Michael TP, Setubal JC, Celton JM, Rees DJ, Williams KP, Holt SH, Ruiz Rojas JJ, Chatterjee M, et al.: The genome of woodland strawberry (Fragaria vesca). Nat Genet 2011, 43:109-116.
• [69]Beyer K, Binder A, Boller T, Collinge M: Identification of potato genes induced during colonization by Phytophthora infestans. Mol Plant Pathol 2001, 2:125-134.
• [70]Baxter L, Tripathy S, Ishaque N, Boot N, Cabral A, Kemen E, Thines M, Ah-Fong A, Anderson R, Badejoko W, Bittner-Eddy P, Boore JL, Chibucos MC, Coates M, Dehal P, Delehaunty K, Dong S, Downton P, Dumas B, Fabro G, Fronick C, Fuerstenberg SI, Fulton L, Gaulin E, Govers F, Hughes L, Humphray S, Jiang RH, Judelson H, Kamoun S, et al.: Signatures of adaptation to obligate biotrophy in the Hyaloperonospora arabidopsidis genome. Science 2010, 330:1549-1551.
• [71]Grouffaud S, van West P, Avrova AO, Birch PR, Whisson SC: Plasmodium falciparum and Hyaloperonospora parasitica effector translocation motifs are functional in Phytophthora infestans. Microbiology 2008, 154:3743-3751.
• [72]Savory EA, Zou C, Adhikari BN, Hamilton JP, Buell CR, Shiu SH, Day B: Alternative splicing of a multi-drug transporter from Pseudoperonospora cubensis generates an RXLR effector protein that elicits a rapid cell death. PLoS One 2012, 7:e34701.
• [73]Yu X, Tang J, Wang Q, Ye W, Tao K, Duan S, Lu C, Yang X, Dong S, Zheng X, Wang Y: The RxLR effector Avh241 from Phytophthora sojae requires plasma membrane localization to induce plant cell death. New Phytol 2012, 196:247-260.
• [74]Cui L, Yin W, Dong S, Wang Y: Analysis of polymorphism and transcription of the effector gene Avr1b in Phytophthora sojae isolates from China virulent to Rps1b. Mol Plant Pathol 2012, 13:114-122.
• [75]Dong S, Kong G, Qutob D, Yu X, Tang J, Kang J, Dai T, Wang H, Gijzen M, Wang Y: The NLP toxin family in Phytophthora sojae includes rapidly evolving groups that lack necrosis-inducing activity. Mol Plant Microbe Interact 2012, 25:896-909.
• [76]Chen X, Shen G, Wang Y, Zheng X, Wang Y: Identification of Phytophthora sojae genes upregulated during the early stage of soybean infection. FEMS Microbiol Lett 2007, 269:280-288.
• [77]Zelaya-Molina LX, Ortega MA, Dorrance AE: Easy and efficient protocol for oomycete DNA extraction suitable for population genetic analysis. Biotechnol Lett 2011, 33:715-720.
• [78]Grabherr MG, Haas BJ, Yassour M, Levin JZ, Thompson DA, Amit I, Adiconis X, Fan L, Raychowdhury R, Zeng Q, Chen Z, Mauceli E, Hacohen N, Gnirke A, Rhind N, di Palma F, Birren BW, Nusbaum C, Lindblad-Toh K, Friedman N, Regev A: Full-length transcriptome assembly from RNA-Seq data without a reference genome. Nat Biotechnol 2011, 29:644-652.
• [79]Pertea G, Huang X, Liang F, Antonescu V, Sultana R, Karamycheva S, Lee Y, White J, Cheung F, Parvizi B, Tsai J, Quackenbush J: TIGR Gene Indices clustering tools (TGICL): a software system for fast clustering of large EST datasets. Bioinformatics 2003, 19:651-652.
• [80]Rice P, Longden I, Bleasby A: EMBOSS: the European molecular biology open software suite. Trends Genet 2000, 16:276-277.
• [81]Bendtsen JD, Nielsen H, von Heijne G, Brunak S: Improved prediction of signal peptides: SignalP 3.0. J Mol Biol 2004, 340:783-795.
• [82]Conesa A, Götz S, García-Gómez JM, Terol J, Talón M, Robles M: Blast2GO: a universal tool for annotation, visualization and analysis in functional genomics research. Bioinformatics 2005, 21:3674-3676.
• [83]Conesa A, Götz S: Blast2GO: A comprehensive suite for functional analysis in plant genomics. Int J Plant Genomics 2008, 2008:619832.
• [84]Moriya Y, Itoh M, Okuda S, Yoshizawa AC, Kanehisa M: KAAS: an automatic genome annotation and pathway reconstruction server. Nucleic Acids Res 2007, 35:W182-W185.
• [85]Altschul SF, Madden TL, Schaffer AA, Zhang J, Zhang Z, Miller W, Lipman DJ: Gapped BLAST and PSI-BLAST: a new generation of protein database search programs. Nucleic Acids Res 1997, 25:3389-3402.
• [86]Enright AJ, Van Dongen S, Ouzounis CA: An efficient algorithm for large-scale detection of protein families. Nucleic Acids Res 2002, 30:1575-1584.
• [87]Eddy SR: Profile hidden Markov models. Bioinformatics 1998, 14:755-763.
• [88]Jones L, Hamilton AJ, Voinnet O, Thomas CL, Maule AJ, Baulcombe DC: RNA-DNA interactions and DNA methylation in post-transcriptional gene silencing. Plant Cell 1999, 11:2291-2301.
• [89]Hall TA: BioEdit: a user-friendly biological sequence alignment editor and analysis program for Windows 95/98/NT. Nucleic Acids Symp Ser 1999, 41:95-98.
• [90]Larkin MA, Blackshields G, Brown NP, Chenna R, McGettigan PA, McWilliam H, Valentin F, Wallace IM, Wilm A, Lopez R, Thompson JD, Gibson TJ, Higgins DG: Clustal W and Clustal X version 2.0. Bioinformatics 2007, 23:2947-2948.
• [91]Yang Z, Wong WS, Nielsen R: Bayes empirical bayes inference of amino acid sites under positive selection. Mol Biol Evol 2005, 22:1107-1118.
• [92]Lévesque CA, Brouwer H, Cano L, Hamilton JP, Holt C, Huitema E, Raffaele S, Robideau GP, Thines M, Win J, Zerillo MM, Beakes GW, Boore JL, Busam D, Dumas B, Ferriera S, Fuerstenberg SI, Gachon CM, Gaulin E, Govers F, Grenville-Briggs L, Horner N, Hostetler J, Jiang RH, Johnson J, Krajaejun T, Lin H, Meijer HJ, Moore B, Morris P, et al.: Genome sequence of the necrotrophic plant pathogen Pythium ultimum reveals original pathogenicity mechanisms and effector repertoire. Genome Biol 2010, 11:R73. BioMed Central Full Text
• [93]Jiang RH, de Bruijn I, Haas BJ, Belmonte R, Löbach L, Christie J, van den Ackerveken G, Bottin A, Bulone V, Díaz-Moreno SM, Dumas B, Fan L, Gaulin E, Govers F, Grenville-Briggs LJ, Horner NR, Levin JZ, Mammella M, Meijer HJ, Morris P, Nusbaum C, Oome S, Phillips AJ, van Rooyen D, Rzeszutek E, Saraiva M, Secombes CJ, Seidl MF, Snel B, Stassen JH, et al.: Distinctive expansion of potential virulence genes in the genome of the oomycete fish pathogen Saprolegnia parasitica. PLoS Genet 2013, 9:e1003272.