Genetics,resistance mechanism,and breeding of powdery mildew resistance in cucumbers(Cucumis sativus L.)

Cucumber is an important vegetable worldwide,and powdery mildew(PM)is a common and serious disease of cucumbers.Breeding disease-resistant cucumber varieties is the most advantageous strategy to control this disease.In recent years,exploration and identification of cucumber PM resistance genes have achieved great advancement,and many genes have been cloned and verified using different methods.However,the resistance mechanism of cucumber PM is still unclear,and many ambiguities need to be elucidated urgently.In this review,we summarized the research advances in PM resistance in cucumbers,including genetic analysis,quantitative trait locus mapping,map-based cloning,transcriptomics,mlo-mediated PM resistance,and mining of noncoding RNAs involved in resistance.Finally,the research directions and the problems that need to be solved in the future were discussed.

Genetic behavior of Triticum aestivum–Dasypyrum villosum translocation chromosomes T6V#4S·6DL and T6V#2S·6AL carrying powdery mildew resistance

T6V#2S·6AL and T6V#4S·6DL translocation chromosomes developed from the cross of wheat and different Dasypyrum villosum accessions have good powdery mildew （PM） resistance, but their pairing and pyramiding behavior remains unclear. Results in this study indicated that the pairing frequency rate of the two differently originated 6VS chromosomes in their F1 hybrid was 18.9% according to genomic in situ hybridization （GISH）; the PM resistance plants in the F2 generation from the cross between T6V#4S·6DL translocation line Pm97033 and its PM susceptible wheat variety Wan7107 was fewer than expected. However, the ratio of the resistant vs. the susceptible plants of 15：1 in the F2 generation derived from the cross between the two translocation lines of T6V#2S·6AL and T6V#4S·6DL fitted well. Plants segregation ratio （homozygous：heterozygous：lacking） revealed by molecular marker for T6V#4S·6DL or T6V#2S·6AL in their F2 populations fitted the expected values of 1：2：1 well, inferring that the pairing of the two alien chromosome arms facilitates the transmission of T6V#4S·6DL from the F1 to the F2 generation. A quadrivalent was also observed in 21% of pollen mother cells （PMCs） of homozygote plants containing the two pairs of translocated chromosomes. The chromosome pairing between 6V#2S and 6V#4S indicates that it will be possible to obtain recombinants and clarify if the PM resistance determinant on one alien chromosome arm is different from that on the other.

Fine mapping of powdery mildew resistance gene PmTm4 in wheat using comparative genomics

Powdery mildew,caused by Blumeria graminis f.sp.tritici,is one of the most severe wheat diseases.Mining powdery mildew resistance genes in wheat cultivars and their appliance in breeding program is a promising way to control this disease.Genetic analysis revealed that a single dominant resistance gene named PmTm4 originated from Chinese wheat line Tangmai 4 confers resistance to prevailing isolates of B.graminis f.sp.tritici isolate E09.Detailed comparative genomics analyses helped to develop closely linked markers to PmTm4 and a fine genetic map was constructed using large F2population,in which PmTm4 was located into a 0.66-c M genetic interval.The orthologous subgenome region of PmTm4in Aegilops tauschii was identified,and two resistance gene analogs（RGA）were characterized from the corresponding sequence scaffolds of Ae.tauschii draft assembly.The closely linked markers and identified Ae.tauschii orthologs in the mapping interval provide an entry point for chromosome landing and map-based cloning of PmTm4.

Comparative genetic mapping revealed powdery mildew resistance gene MlWE4 derived from wild emmer is located in same genomic region of Pm36 and Ml3D232 on chromosome 5BL

Powdery mildew, caused by Blumeria graminis f. sp. tritici, is one of the most devastating wheat diseases. Wild emmer wheat（Triticum turgidum ssp. dicoccoides） is a promising source of disease resistance for wheat. A powdery mildew resistance gene conferring resistance to B. graminis f. sp. tritici isolate E09, originating from wild emmer wheat, has been transferred into the hexaploid wheat line WE4 through crossing and backcrossing. Genetic analyses indicated that the powdery mildew resistance was controlled by a single dominant gene, temporarily designated Ml WE4. By mean of comparative genomics and bulked segregant analysis, a genetic linkage map of Ml WE4 was constructed, and Ml WE4 was mapped on the distal region of chromosome arm 5BL. Comparative genetic linkage maps showed that genes Ml WE4, Pm36 and Ml3D232 were co-segregated with markers XBD37670 and XBD37680, indicating they are likely the same gene or alleles in the same locus. The co-segregated markers provide a starting point for chromosome landing and map-based cloning of Ml WE4, Pm36 and Ml3D232.

Characterization of RAPD Markers,and the RFLP Marker Linked to Powdery Mildew Resistance Gene Derived from Different Accessions of H.villosa

The analysis was carried out on performance of the resistance gene from Haynaldia villosa accession of the former Soviet Union to different isolates of Bluemerie graminis. Polymorphisms were revealed between 6D/6V substitution line Pm930640 and its pedigree parents using five RAPD markers of OPAN031700, OPAI01700, OPAL03750, OPAD07480 and OPAG15580 screened out from 120 random 10-mers primers. Three RAPD markers of OPAN03, OPAI01 and OPAL03 were linked with the resistance gene by analysis of F2 population of Chancellor×Pm930640. Analysis of 29 wheat lines including part of lines conferring the known genes from Pm1 to Pm20 respectively, lines conferring resistance gene from two H. villosa accessions and the related wheat parents, were analyzed and the results showed that these markers not only linked to the gene resistant to powdery mildew from H. villosa, but also detected different genetic backgrounds. OPAL03750 can be used as the marker to distinguish the different resistant lines from two H. villosa accessions because it was only observed in the materials from H. villosa of the former Soviet Union. RFLP analysis also showed the polymorphisms between two H. villosa accessions and their derived resistant lines.

A single-nucleotide substitution in the leucinerich-repeat-only gene CsLRR1 confers powdery mildew resistance in cucumber

Dear Editor,A variant with single substitution in the LRR-only protein CsLRR1,which regulates cucumber resistance to powdery mildew through a salicylic-acid-mediated pathway,provides a major source of resistance for cucumber breeding.Cucumber powdery mildew(PM),mainly caused by the biotrophic pathogen Podosphaera xanthii(synonym Podosphaera fusca),poses a serious threat to cucumber crops globally,leading to substantial reductions in fruit yield and quality(Martinez-Cruz et al.,2017).

Orthologous genesPm12andPm21from twowild relatives of wheat show evolutionary conservation but divergent powdery mildew resistance

Wheat powdery mildew,caused by Blumeria graminis f.sp.tritici(Bgt),is a devastating disease that threatens wheat production worldwide.Pm12,which originated from Aegilops speltoides,a wild relative of wheat,confers strong resistance to powdery mildew and therefore has potential use in wheat breeding.Using susceptible mutants induced by gamma irradiation,we physically mapped and isolated Pm12 and showed it to be orthologous to Pm21 from Dasypyrum villosum,also a wild relative of wheat.The resistance function of Pm12 was validated via ethyl methanesulfonatemutagenesis,virus-induced gene silencing,and stable genetic transformation.Evolutionary analysis indicates that the Pm12/Pm21 loci in wheat species are relatively conserved but dynamic.Here,we demonstrated that the two orthologous genes,Pm12 and Pm21,possess differential resistance against the same set of Bgt isolates.Overexpression of the coiledcoil domains of both PM12 and PM21 induces cell death in Nicotiana benthamiana leaves.However,their full-length forms display different cell death-inducing activities caused by their distinct intramolecular interactions.Cloning of Pm12 will facilitate its application in wheat breeding programs.This study also gives new insight into two orthologous resistance genes,Pm12 and Pm21,which show different race specificities and intramolecular interaction patterns.

Genome Analysis in Wheat Breeding for Disease Resistance

A brief review on the development of wheat germplasm with introduced powdery mildew and scab resistance from Haynaldia villosa Sch. and Leymus racemosus Lam., Roegneria ciliaris (Trin.) Nevski as well as R. kamoji C. Koch respectively was made. In the course of germplasm development, genome analysis by means of chromosome banding, genomic in situ hybridization (GISH) or fluorescence in situ hybridization (FISH), molecular markers, particularly restriction fragment length polymorphism (RFLP) coupled with aneuploid analysis was employed for the purpose of improving breeding efficiency. Potential use of such germplasm in wheat breeding practice, basic studies and some related problems were also discussed.

Introgression of Resistance to Powdery Mildew Conferred by Chromosome 2R by Crossing Wheat Nullisomic 2D with Rye

Using the nulUsomic back-cross procedure, four wheat-rye chromosome substitution 2R （2D） lines with different agronomic performance, designated WR02-145-1, WR01-145-2, WR02-145-3, and WR02-145-4, were produced from a cross between 2D nullisomic wheat （Triticum aestivum L. cv. ＂Xiaoyan 6＂） and rye （Secale cereale L. cv. ＂German White＂）. The chromosomal constitution of 2n=42=21 in WR02-145 lines was confirmed by cytological and molecular cytogenetic methods. Using genomic in situ hybridization on root tip chromosome preparations, a pair of intact rye chromosomes was detected in the WR02-145 lines. PCR using chromosome-specific primers confirmed the presence of 2R chromosomes of rye in these wheat-rye lines, indicating that WR02o145 lines are disomic chromosome substitution lines 2R （2D）. The WR02-145 lines are resistant to the powdery mildew （Erysiphe graminis DC. f. sp. tritici E. Marchal） isolates prevalent in northern China and may possess gene（s） for resistance to powdery mildew, which differ from the previously identified Pm7gene located on chromosome 2RL. The newly developed ＂Xiaoyan 6＂- ＂German White＂ 2R （2D） chromosome substitution lines are genetically stable, show desirable agronomic traits, and are expected to be useful in wheat improvement.

Genome-wide identification and evolutionary analysis of MLO gene family in Rosaceae plants

Mutants lacking wild-type MLO(Mildew resistance Locus O)proteins show broad-spectrum resistance to the powdery mildew fungus,and dysregulated cell death control,with spontaneous cell death in response to developmental or abiotic stimuli.In order to understand the evolution and divergence patterns of the MLO gene family in Rosaceae plants,we analysed systematically genome-wide data from Fragaria vesca,Prunus persica,Prunus mume,Malus domestica,Pyrus bretschneideri and Rubus occidentalis based on bioinformatics methods.Using three phylogenetic methods(the neighbour-joining,maximum likelihood,and Bayesian methods),we identified 117 MLO genes from 6 Rosaceae species.The results of all three phylogenetic analysis methods supported that these genes were divided into six clades.Conserved motif analysis found that only motif 2 was present in all MLO proteins and had 3 nearly invariant amino acid residues.The findings indicated that motif 2 might be shared by the MLO gene family.The structural features of these genes showed large variations in sequence length among different species,although the lengths and the numbers of exons exhibited high degrees of similarity.Selective pressure analysis showed extremely significant differences in all 6 clades,with 2,1,and 1 site(s)under significant positive selection detected in clades III,IV,and VI,respectively.These positive selection sites were important driving forces for the promotion of the functional differentiation of the MLO genes.Functional divergence analysis showed that the significantly divergent sites were located within the domains of the MLO genes.Functional distance analysis showed that the clade V had more conservative functions and might have retained more original functions during the evolutionary process.However,clade I may have undergone extensive altered functional constraints as a specialised functional role.Moreover,the most original function of the MLO genes in Rosaceae could be related to the evolution of their resistance to powdery mildew,which then gradually evolved into functions such as the regulation of flower development,the control of root morphology,and seed evolution due to the different evolutionary rates after gene duplication.These results provide a theoretical basis for further studies of the molecular evolutionary patterns of the plant MLO gene family.

Characterization of Three Novel Wheat LThinopyrum intermedium Addition Lines with Novel Storage Protein Subunits and Resistance to Both Powdery Mildew and Stripe Rust

Wide hybridization is an effective approach for enhancing the resistance of bread wheat （Triticum aestivum L.） to biotic and abiotic stresses by introducing favorable alien genes （Sepsi et al., 2008）. Wheatgrass, Thinopyrum intermedium （Host） Barkworth ＆ D.R. Dewey or Agropyron intermedium （Host） Beauvoir （2n = 42; genome formula JJjSjSstst）, is a perennial species in the tribe Triticeae and an important source of wheat improvement for biotic and abiotic stress resistance and quality-related traits, such as high grain protein concentration （Chen et al., 1998; 2001; 2003; Han et al., 2004; Li and Wang, 2009）. In addition, the ready crossing ability of wheatgrass with various Triticum species has made it popular in germ- plasm development.