Establishment and application of gene tagging linked to rice blast resistance gene Pi-d2.
2
Citation
0
Reference
20
Related Paper
Citation Trend
Abstract:
The establishment of molecular marker of rice blast resistance genes was a powerful strategy in resistance to rice blast breeding.In the present study,seventy-one strains in Digu variety was used to check the regional rice blast resistance in Guangxi.A gene tagging M-Pid2 was amplified by the primers designed with the sequence of resistance gene itself,using a base difference between Pi-d2 gene in Digu variety and Guanghui 998 allele.The validity of the gene tag was tested in a F2 generation derived from Digu(a resistant variety including Pi-d2) and Guanghui 998(a susceptible restore line) and a group of germplasm containing 62 rice germplasm.The resistance spectrum was detected by inoculating Pi-d2 gene sponsor with 71 rice blast pathogen strains collected from Guangxi.The resistance spectrum of all materials were also detected in natural disease nursery located in Cenxi county.The results showed that the resistance indicated that Pi-d2 gene tag was highly consistent with that indicated under field detection.Furthermore,PCR analysis indicated that,among the 62 rice germplasm,a specific band of 629 bp was amplified only in Digu variety.It was concluded that rice blast resistance gene Pi-d2 was the main-effect blast resistance genes of Digu rice.and the gene tag could be credibly used in marker assisted selection and germplasm differentiation.Keywords:
Germ plasm
R gene
Cite
The establishment of molecular markers of rice blast resistance genes was a powerful strategy in rice blast resistance breeding.In present study,PCR markers for four blast resistance genes(Pi9、Pi2、Pi5 and Pita) were screened by the primers designed based on the sequences of four blast genes and their variances in Nipponbare.These markers were tested in 24 rice lines carrying a single blast resistance gene and 48 rice parents.Rice blast resistances of 48 rice parents were also tested with rice blast pathogens ZB1 and ZB13 collected from Guangxi.The results showed that these markers were effective to identify blast resistance genes.The marker M-Pi9 showed a specific amplified band in the lines carrying the gene Pi9 or Piz among the 24 rice lines with a single blast resistance gene;M-Pi2 showed specificity in the lines with gene Pi2 or Pizt;M-Pi5 showed specificity in three lines with genes Pii,Pi3 or Pi5;and M-Pita showed specificity in the lines with the gene Pita or Pita2.Furthermore,we found that the rice parents with the specific marker M-Pi2 showed resistance to rice blast pathogen ZB1,while that with four markers(M-Pi9,M-Pi2,M-Pi5 and M-Pita) showed resistance to rice blast pathogen ZB13.
Molecular marker
Cite
Citations (4)
Abstract Background As rice ( Oryza sativa ) is the staple food of more than half the world’s population, rice production contributes greatly to global food security. Rice blast caused by the fungus Magnaporthe oryzae ( M. oryzae ) is a devastating disease that affects rice yields and grain quality, resulting in substantial economic losses annually. Because the fungus evolves rapidly, the resistance conferred by most the single blast-resistance genes is broken after a few years of intensive agricultural use. Therefore, effective resistance breeding in rice requires continual enrichment of the reservoir of resistance genes, alleles, or QTLs. Seed banks represent a rich source of genetic diversity; however, they have not been extensively used to identify novel genes and alleles. Results We carried out a large-scale screen for novel blast-resistance alleles in 1883 rice varieties from major rice-producing areas across China. Of these, 361 varieties showed at least moderate resistance to natural infection by rice blast at rice blast nurseries in Enshi and Yichang, Hubei Province. We used sequence-based allele mining to amplify and sequence the allelic variants of the major rice blast-resistance genes at the Pi2 / Pi9 locus of chromosome 6 from the 361 blast-resistant varieties, and the full-length coding region of this gene could be amplified from 107 varieties. Thirteen novel Pi9 alleles (named Pi9 -Type1 to Pi9 -Type13) were identified in these 107 varieties based on comparison to the Pi9 referenced sequence. Based on the sequencing results, the Pi2/Pi9 locus of the 107 varieties was divided into 15 genotypes (including three different genotypes of Pi9 -Type5). Fifteen varieties, each representing one genotype, were evaluated for resistance to 34 M. oryzae isolates. The alleles from seven varieties with the highest resistance and widest resistance spectra were selected for transformation into the susceptible variety J23B to construct near-isogenic lines (NILs). These NILs showed resistance in a field test in Enshi and Yichang, indicating that the seven novel rice blast-resistance tandem-repeat regions at the Pi2/Pi9 locus of chromosome 6 could potentially serve as a genetic resource for molecular breeding of resistance to rice blast. Conclusions The thirteen novel Pi9 alleles identified in this study expand the list of available of blast-resistance alleles. Seven tandem-repeat regions of the Pi2/Pi9 locus from different donors were characterized as broad-spectrum rice blast-resistance fragments; these donors enrich the genetic resources available for rice blast-resistance breeding programs.
Pyricularia
Cite
Citations (16)
The rice Pi2/9 locus harbors multiple resistance (R) genes each controlling broad-spectrum resistance against diverse isolates of Magnaporthe oryzae, a fungal pathogen causing devastating blast disease to rice. Identification of more resistance germplasm containing novel R genes at or tightly linked to the Pi2/9 locus would promote breeding of resistance rice cultivars.In this study, we aim to identify resistant germplasm containing novel R genes at or tightly linked to the Pi2/9 locus using a molecular marker, designated as Pi2/9-RH (Pi2/9 resistant haplotype), developed from the 5' portion of the Pi2 sequence which was conserved only in the rice lines containing functional Pi2/9 alleles. DNA analysis using Pi2/9-RH identified 24 positive lines in 55 shortlisted landraces which showed resistance to 4 rice blast isolates. Analysis of partial sequences of the full-length cDNAs of Pi2/9 homologues resulted in the clustering of these 24 lines into 5 haplotypes each containing different Pi2/9 homologues which were designated as Pi2/9-A5, -A15, -A42, -A53, and -A54. Interestingly, Pi2/9-A5 and Pi2/9-A54 are identical to Piz-t and Pi2, respectively. To validate the association of other three novel Pi2/9 homologues with the blast resistance, monogenic lines at BC3F3 generation were generated by marker assisted backcrossing (MABC). Resistance assessment of the derived monogenic lines in both the greenhouse and the field hotspot indicated that they all controlled broad-spectrum resistance against rice blast. Moreover, genetic analysis revealed that the blast resistance of these three monogenic lines was co-segregated with Pi2/9-RH, suggesting that the Pi2/9 locus or tightly linked loci could be responsible for the resistance.The newly developed marker Pi2/9-RH could be used as a potentially diagnostic marker for the quick identification of resistant donors containing functional Pi2/9 alleles or unknown linked R genes. The three new monogenic lines containing the Pi2/9 introgression segment could be used as valuable materials for disease assessment and resistance donors in breeding program.
Germ plasm
Identification
Cite
Citations (9)
Pyricularia
Inbred strain
Major gene
Genetic linkage
Cite
Citations (24)
Germ plasm
Introgression
Magnaporthe
Cite
Citations (13)
Gene pyramiding is an effective approach for the improvement of rice varieties with durable resis-tance to blast by crossing among some blast-resistant rice varieties.To blast-resistant rice resource,the foun-dation of marker-assisted selection(MAS) is resistance genotype identification with molecular markers.Some rice cultivars resistant to rice blast were selected by certifying with rice blast plot.Meanwhile,specific molecular markers were used to test the blast resistance genes Pi9,Pita,Pib and Pikm in the cultivars and the database contained resistant gene were established initially.Furtherly,the correlation between resistant gene and the disease reactions was discussed,and the result indicated that the durable blast-resistant ability would be improved when Pi9 was a major gene polymerized with Pita,Pib.
Blast disease
Molecular marker
Cite
Citations (4)
Pyricularia
Blast disease
R gene
Cite
Citations (10)
Summary Because of the frequent breakdown of major resistance ( R ) genes, identification of new partial R genes against rice blast disease is an important goal of rice breeding. In this study, we used a core collection of the Rice Diversity Panel II (C‐RDP‐II), which contains 584 rice accessions and are genotyped with 700 000 single‐nucleotide polymorphism (SNP) markers. The C‐RDP‐II accessions were inoculated with three blast strains collected from different rice‐growing regions in China. Genome‐wide association study identified 27 loci associated with rice blast resistance (LABRs). Among them, 22 LABRs were not associated with any known blast R genes or QTLs. Interestingly, a nucleotide‐binding site leucine‐rich repeat (NLR) gene cluster exists in the LABR12 region on chromosome 4. One of the NLR genes is highly conserved in multiple partially resistant rice cultivars, and its expression is significantly up‐regulated at the early stages of rice blast infection. Knockout of this gene via CRISPR‐Cas9 in transgenic plants partially reduced blast resistance to four blast strains. The identification of this new non‐strain specific partial R gene, tentatively named rice blast Partial Resistance gene 1 ( PiPR1 ), provides genetic material that will be useful for understanding the partial resistance mechanism and for breeding durably resistant cultivars against blast disease of rice.
Oryza
Cite
Citations (49)
In order to discover,identify and apply the new broad-spectrum resistant genes,this research had identified one new rice blast broad-spectrum resistant allele. In this study,according to the sequence of rice blast broad-spectrum resistance recessive allele pi21,molecular marker Pi21-1 was designed and amplified using wide compatibility rice variety 02428. Sequencing analysis showed new mutation type,and rice blast vaccination identification resulted in immunity. Thus,a new rice blast broad-spectrum resistance allele pi21 t had been identified. These results provided new resistant resources for rice blast broad-spectrum resistant breeding. Meanwhile,genotype of target variety could be determined rapidly using newly designed marker. All these accelerate the process of rice blast broad-spectrum resistant breeding.
Broad spectrum
Cite
Citations (0)
The phenotype of broad-spectrum rice blast resistance variety Yunyin was analyzed and identified by inoculated with rice blast fungi RB01-RB22 of known avirulence genes as pathogen in vitro.The results showed that Yunyin was resistant to all rice blast fungi except RB17 and RB20.Analyzing the widely used 24 resistance genes in production at present,the results showed that Yunyin may contain new blast resistance genes besides these genes,and there were able to be applied for rice breeding and production.In addition,in this study,we constructed near-isogenic lines with LTH as genetic background and contained blast-resistant gene to blast isolate SiChuan 43 by successive backcrossing combining marker-assisted selection with artificial inoculation.These works would be beneficial to the genetics study and application of this blast-resistance gene.
Genetic Analysis
Cite
Citations (2)