Avirulence of Magnaporthe grisea isolate CHL346 on rice cultivar GA25 was studied with 242 ascospore progenies derived from the cross CHL346 x CHL42. Segregation analysis of the avirulence in the progeny population was in agreement with the existence of a single avirulence (Avr) gene, designated as AvrPi15. For mapping the Avr gene, we developed a total of 121 microsatellite DNA markers [simple sequence repeat (SSR)], which evenly distributed in the whole-genome of M. grisea through bioinformatics analysis (BIA) using the publicly available sequence. Linkage analysis of the AvrPi15 gene with these SSR markers showed that six markers on chromosome 6, MS6-1, MS6-2, MS6-3, MS6-7, MS6-8 and MS6-10, were linked to the AvrPi15 locus. To further define the chromosomal location of the AvrPi15 locus, two additional markers, MS6-17 and STS6-6, which were developed based on the sequences of telomeric region 11 (TEL11), were subjected to linkage analysis. The results showed that MS6-17 and STS6-6 were associated with the locus by 3.3 and 0.8 cM, respectively. To finely map the Avr gene, two additional candidate avirulence gene (CAG) markers, CAG6-1 and CAG6-2, were developed based on the gene annotation of the sequence of TEL 11. Linkage analysis of the Avr gene with these two markers revealed that both of them completely cosegregated with the AvrPi15 locus. Finally, this locus was physically mapped into approximately 7.2-kb interval of the TEL11 by BIA using these sequence-ready markers. This is the key step toward positional cloning of the AvrPi15 gene.
The results indicated the powerful ability of MGP1 as a biological agent.
Magnaporthe oryzae is a model for plant pathogenic filamentous fungi. We have assembled a simple sequence repeat (SSR)-based physical map of the species, using in silico sequence data. A set of 120 SSR markers was developed from the genomic sequence of the reference isolate 70-15. These markers were readily amplified from the genomic DNA of other isolates, and high levels of allelic variation characterised the parental isolates of the two crosses tested. All the markers were locatable to one of the seven M. oryzae chromosomes. An SSR-based physical in silico map was constructed, and pre-existing SSR and RFLP loci were integrated into the map, along with 23 Avr (avirulence) genes and two other genes of importance to the plant/pathogen interaction. This map provides a platform for population genetics and functional genomics studies in the model pathogen, and even in other evolutionally related pathogens.Magnaporthe oryzae, simple sequence repeat (SSR), reference sequence, physical map, in silico mapping
Rice blast, caused by the fungal pathogenMagnaporthe oryzae, is one of the most devastating crop diseases worldwide. The avirulence gene corresponding to rice blast resistance gene Pi7 in field isolate CHL346 was inherited as a single gene, designated AvrPi7, in a segregating population consisting of 189 ascospore progenies derived from a cross between field isolates CHL346 and CHL42. In order to determine the chromosomal location of the AvrPi7 locus, a total of 121 simple sequence repeat (SSR) markers were developed based on the whole-genome sequence of reference isolate 70-15 of M. oryzae. Linkage analysis of the locus with these SSR markers showed that eight SSR markers on chromosome 1 were linked to the locus, among which the closest flanking markers MS1-9 and MS1-15 were 3.2 and 16.4 cM from the locus, respectively. For fine mapping, additional PCR-based makers including eight SSR markers and three candidate avirulence gene (CAG) markers were developed in the region flanking both markers. The AvrPi7 locus was genetically delimited within a 1.6-cM region flanked by markers MS1-21 and MS1-22, and co-segregated with the marker CAG2. To construct a physical map of the AvrPi7 locus, molecular markers linked to the Avr gene were mapped on the supercontigs of the reference isolate 70-15 through bioinformation analysis (BIA). Consequently, the AvrPi7 locus was delimited to a 75-kb interval flanked by markers MS1-21 and MS1-22 based on the reference sequence. Merodiploids observed in this study are also discussed.
aThe first two authors contributed equally to this work. AbstractThe ability of Rhizoctonia solani AG-1 IA, the causal agent of rice sheath blight, to survive in diseased rice straw and as sclerotia and mycelia was investigated. After storage for 10 months at 4°C, 25°C and non-air-conditioned natural room temperature (NRT, temperature range from 6°C to 35°C), sclerotia placed inside a desiccator, soaked in sterile water or immersed in wet paddy soil were viable. In contrast, only 15% of sclerotia in dry paddy soil survived. Survival of mycelia was severely affected by temperature and humidity. After 10 months in a desiccator at 4°C, 55% of mycelia samples could survive, whereas at 25°C and NRT, mycelial samples survived for only 7 and 5 months, respectively. However, mycelia stored in sterile water at constant temperatures (4°C or 25°C) survived for 10 months. A certain amount of UV radiation had no obvious effect on the survival of sclerotia or mycelia. The survival rate of the fungus in diseased rice straw stored for 16 months could reach 100% at 4°C, 50% at 25°C and 35% at NRT. The survival rates of the pathogen in diseased rice straw buried in dry, wet and flooded paddy soils after 10-month storage at NRT were 75, 100 and 100%, respectively, indicating that soil humidity is a crucial factor for the survival of this fungus.
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