| The pathogenicity variation(new virulent races producing)of Puccinia striiformis and its spreading are the primary factors leading to the breakdown of resistant stripe rust genes in the host.Somatic recombination and sexual recombination had been hypothesized as the important reason for the variation of Puccinia striiformis f.sp.tritici(Pst)with limited evidence.Developing somatic recombination and sexual recombination isolates under controlled experimental conditions and characterizing their virulence and molecular phenotypes with reliable markers that benefit to reveal the variation mechanism of Pst and provide theoretical guidance for prediction of new race as well as breeding and distribution of resistance cultivars.In the study of somatic recombination,We characterize the potential somatic recombinant isolates(R isolates)using a new set of Yr single-gene differentials,and larger numbers of the co-dominant simple sequence repeat(SSR)and secrete ptrotein single-nucleotide polymorphism(SP-SNP)markers.Results are shown below.1.Based on the results of the new set of 18 Yr single-gene lines and 141 molecular markers(51 SSR and 90 SP-SNP)of parental and their progeny isolates,67 of the 68 progeny isolates in 9 combination are thought to be recombinants.2.In the virulence tests,virulence phenotype recombination was observed in progeny isolates and there are possible four types of recombination,those are losing virulence or partial virulence(gaining avirulence),gaining partial virulence or virulence(losing avirulence),recombined virulence,recombined virulence and avirulence.In somatic recombination,the segregation and recombination of virulence/avirulence in progeny isolates opens a new avenue to study the pathogen population and the genetics of pathogen.3.In the three combinations of Pst and Puccinia striiformis f.sp.tritici(Psh),16 progeny isolates were considered to be possible recombinants based on virulence tests and 15 of them were thought to be recombinants though further molecular marker analysis.It indicated that recombination can occur between Pst and Psh under controlled experimental conditions.However,based on the number of R isolates,it appears that R isolates may be produced more easily between isolates of a same forma specialis than between different formae speciales.4.Our marker data indicate that almost all of the progeny isolates were produced through somatic recombination including the processes of cell fusion,nuclear re-assortment(somatic hybridization),chromosomal re-assortment,and crossover.This study is the first to use a large number of DNA-based markers to characterize somatic recombination in P.striiformis and prove that somatic recombination can occur between P,striiformis isolates under controlled greenhouse conditions.Somatic recombination is a mechanism for P.striiformis variation under controlled conditions.In sexual recombination,we developed a progeny population comprised of 117 unique isolates through selfing Pst isolate PSTv-4 on barberry plants.The selfing population(including isolates PSTv-4 and its progeny isolates)were analyzed by 35 Yr single-gene lines and molecular markers,and preliminary linkage maps were construction based on the results of virulence and molecular phenotypes.1.The parental isolate and all progeny isolates were all avirulent to resistance genes Yr5,Yr10,Yr15,Yr24,Yr32,YrTrl,Yr26,YrCV,YrTre,Yr45,Yr53,and Yr64,but virulent to Yr1,Yr6,Yr9,YrSP,YrTye,Yr2,Yr21,Yr25,Yr28,Yr29,Yr31,YrA,YrAvs,and Yr65,indicating the parental isolate was homozygous in avirulence to the former group of resistance genes,and homozygous in virulence to the latter group of resistance genes.Segregation were observed for avirulence to Yr7,Yr8,Yr43,Yr44,YrExp2 and for virulence to Yr17,Yr27,Yr35,Yr41,that mean the parental isolate was heterozygous in avirulence to Yr7,Yr8,Yr43,Yr44,YrExp2 and virulence to Yr17,Yr27,Yr35,Yr41.2.Counting the number of avirulence and virulence isolates on segregation differentials and using Chi-square test for goodness of fit of the observed isolates in avirulent and virulent classes to theoretical ratios,we found that the parental isolate had one dominant gene each for avirulence to Yr1,Yr43,Yr44 and YrExp2;one recessive gene for avirulence to Yrl7 and Yr27;and two dominant gene for avirulence Yr8;and two recessive genes for avirulence to Yr35 and Yr41.3.From 117 isolates,38 virulence patterns were identified.Of the 117 progeny isolates,there were 40 progeny isolates(34.19 of progeny isolates)which had a wider virulence formulae than that of parental isolate.Even more,five isolates were virulent on all the 9 segregation differentials.The results showed that through sexual reproduction,a single isolates can generate a large number of races with different virulences,and more virulent races could be produced.4.In the total of 342 pairs of SSR primers and 189 pairs of SP-SNP primers,53 of them were available for drawing linkage maps,including 24 pairs of SSR primers and 29 pairs of SP-SNP primers.Based on the virulence and molecular phenotypes,9 linkage groups were obtained by software of Mapmaker/EXP3.0 and MapDaw2.1.The linkage groups covering a total of 711.2 cM and the average distance of two loci was about 19.8 cM.9 avirulence loci were distributed in three linkage groups.Among them,avirulence genes Avr7,Avr44,AvrExp2 and Avr43 were clustered tightly,while avr 35,avr 27,avr 41 and avrl 7 clustered together.However,none of molecular marker in this study linked with them.Avr8 linked to 8 molecular markers and the nearest genetic distance(36.6 cM)to Avr8 was SSR marker PstP033.The pathogenicity variation of selfing population,the homozygous or heterozygous of avirulence/virulence and the genetic traits of avirulence were discussed,in the same time,we construct a preliminary linkage maps.These results can provide the basis to the research of sexual recombination and virulence variation,and for the further clone of avirulence genes.Somatic recombination and Sexual recombination can be the two important ways of genetic variation in Pst under controlled conditions. |
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