Evaluation Of Major Potato Varieties In Northwestern China For Late Blight Resistance And Identification Of Candidate Core RXLR Effector Genes From Phytophthora Infestans

Potato is an important staple crop ranking fourth in global production after maize,rice and wheat.It plays important role in global food security.Late blight,caused by oomycete pathogen Phytophthora infestans,is the most destructive disease of potato and seriously threaten potato's sustainable production.It is notorious for causing the “Irish famine” disaster in 1840 s.Applying resistance potato cultivars is the most effective and economic and environmental friendly strategy to control late blight diseases.However,P.infestans can rapidly variaty and escape from being recognized by potato resistance genes and causes the outbreaks and epidemics of late blight constantly,which makes itself called “Resistance gene destroyer”.Northwest area of China is one of the main potato production regions,but the potato production has been constantly threatened by late blight disease.Currently,decrease the using of chemical agents and increase their controlling effect are becoming our pursuing goal for green agriculture.Under such kind of background,how to effectively and durably control late blight disease and breed durable resistant potato cultivars are emerging to be our essential purpose.Because little information is available about the resistance performance of major potato varieties in northwest area of China,as well as major varieties' resistance genetic background,which make it hard to deploy resistance varieties properly and result in the vulnerable of potato production.Thus,we evaluated the field resistance performance of 101 varieties.And through using 10 known avirulence RXLR effector genes as tools,we also dissected the resistance gene(s)obtained by 20 dominant varieties.Meanwhile,these potato resistant genes targeting to conserved core RXLR effector genes are potentially more durable.In order to identify the core RXLR effector genes expressing in early infection stage,we selected six P.infestans strains with diverse genetic background to prepare the 12 hpi(hours post inoculation)infected potato leaf materials and then perform the RNA-seq sequencing.A group of candidate core RXLR effector(CRE)genes were identified from the sequencing data.And then part of candidate CRE genes were selected to examine their potential virulence function using Agrobacterium tumefaciens-mediated transient expression on Nicotiana benthamiana and Solanum tuberosum.The results are summarized as follows:1.Field resistance evaluation showed that 19 were high resistant varieties,while most varieties performed moderate resistant to high susceptible to P.infestans.By using 10 known avirulence genes as tools,we dissected the resistance gene(s)obtained by 20 dominant varieties.The result showed that these varieties contain 0 to 10 resistance genes matching to cloned Avr gens,with an average of five genes in each.When the field epidemic population was dominated by high virulence strains,the correlation analysis between field disease index(AUDPC)and resistance gene number revealed significant negative correlation(R = 0.70**,P < 0.05),which suggested that more resistance genes contribute higher resistance level of potato to late blight.2.By Illumina high throughput RNA-seq sequencing,46 G data was produced from 6 samples.Among them,2,337,987(1%)reads belong to P.infestans.After gene assembling,11,911 genes were detected expressing in six strains,with an average of 7,000 genes in each.Among the detected genes,245 were RXLR effectors genes and averagely 160 genes were detected in each strain.Expression profiling indicated that the expression level of RXLR effector genes was 2-to 3-fold higher than that of whole transcriptome,suggesting that many RXLR effector genes are induced during early infection stage.A total of 468 SNPs was detected in 128 RXLR effector genes,including 183 synonymous and 266 nonsynonymous substitutions,revealing that RXLR genes undergo positive selection pressure resulting in higher polymorphism.Based on the parameters of sequence conservation and infection induced expression,23 candidate RXLR effector genes were determined.After the phylogenetic analysis,those genes were finally designated as 18 CRE genes.Nine CREs were selected to test their virulence contribution and found all of them can enhance the colonization of P.infestans.Virulence function of four CREs were performed on N.benthamiana and S.tuberosum using Agrobacterium tumefaciens-mediated transient expression assay.The results displayed that the tested RXLR genes are capable of suppressing PTI(PAMP triggered immunity)and ETI(Effector trigged immunity)defense response induced by BAX,INF1,NIP,Avh241,and Avh238.Those tests suggested that CREs contribute to virulence by suppressing plant defense response during early infection stage.