Characterization of Rice Blast Isolates by the Differential System and their Application for Mapping a Resistance Gene,Pi19(t)

To facilitate resistance gene characterization in the present study, the pathogenicities of newly collected blast isolates from rice fields in the Philippines were characterized using international blast differential varieties consisting of 31 monogenic lines that target 24 resistance genes. To classify and designate the blast isolates, we used a new international blast designation system, which has been proposed as a suitable naming system for comparing blast races among different studies. A total of 23 rice blast isolates collected from the Philippines were classified into 16 pathotypes, which showed reaction patterns different from those seen in the standard isolates. Among the blast pathotypes, 11 had differentiating ability for four Pik alleles (Pik, Pik‐m, Pik‐h, and Pik‐p) and Pi1, whereas the standard blast isolates from the Philippines were not able to differentiate these genes. In addition, several blast isolates were avirulent to IRBLt‐K59, IRBL19‐A, and Lijiangxintuanheigu, although the standard differential blast isolates were virulent to these lines. Moreover, two blast isolates were virulent to a monogenic line, IRBL9‐W, which harbours Pi9 and was resistant to all standard differential blast isolates. By using the isolates avirulent to IRBL19‐A, Pi19(t) was successfully mapped in the centromeric region on chromosome 12 with simple sequence repeat markers RM27937 and RM1337. These markers are useful for marker‐assisted Pi19(t) introgression worldwide.     

A novel gene, <Emphasis Type="Italic">Pi40(t)</Emphasis>, linked to the DNA markers derived from NBS-LRR motifs confers broad spectrum of blast resistance in rice

Rice blast disease caused by Magnaporthe grisea is a continuous threat to stable rice production worldwide. In a modernized agricultural system, the development of varieties with broad-spectrum and durable resistance to blast disease is essential for increased rice production and sustainability. In this study, a new gene is identified in the introgression line IR65482-4-136-2-2 that has inherited the resistance gene from an EE genome wild Oryza species, O. australiensis (Acc. 100882). Genetic and molecular analysis localized a major resistance gene, Pi40(t), on the short arm of chromosome 6, where four blast resistance genes (Piz, Piz-5, Piz-t, and Pi9) were also identified, flanked by the markers S2539 and RM3330. Through e-Landing, 14 BAC/PAC clones within the 1.81-Mb equivalent virtual contig were identified on Rice Pseudomolecule3. Highly stringent primer sets designed for 6 NBS-LRR motifs located within PAC clone P0649C11 facilitated high-resolution mapping of the new resistance gene, Pi40(t). Following association analysis and detailed haplotyping approaches, a DNA marker, 9871.T7E2b, was identified to be linked to the Pi40(t) gene at the 70 Kb chromosomal region, and differentiated the Pi40(t) gene from the LTH monogenic differential lines possessing genes Piz, Piz-5, Piz-t, and Pi-9. Pi40(t) was validated using the most virulent isolates of Korea as well as the Philippines, suggesting a broad spectrum for the resistance gene. Marker-assisted selection (MAS) and pathotyping of BC progenies having two japonica cultivar genetic backgrounds further supported the potential of the resistance gene in rice breeding. Our study based on new gene identification strategies provides insight into novel genetic resources for blast resistance as well as future studies on cloning and functional analysis of a blast resistance gene useful for rice improvement.     

Polymorphism analysis of genomic regions associated with broad-spectrum effective blast resistance genes for marker development in rice

Cultivated European rice germplasm is generally characterized by moderate to high sensitivity to blast, and blast resistance is therefore one of the most important traits to improve in rice breeding. We collected a panel of 25 rice genotypes containing 13 broad range rice resistance genes that are commonly used in breeding programs around the world: Pi1, Pi2, Pi5, Pi7, Pi9, Pi33, Pib, Pik, Pik-p, Pita, Pita ² , Piz and Piz-t. The efficiency of the selected Pi genes towards Italian blast pathotypes was tested via artificial inoculation and under natural field infection conditions. To characterize haplotypes present in the chromosomal regions of the blast resistance genes, a polymorphism search was conducted in the sequence regions adjacent to the blast resistance by examining DNA from the Pi gene donors with a panel of 5–7 potential receivers (cultivated European rice genotypes). Seven InDel and 8 presence/absence polymorphisms were directly detected by gel analysis after DNA amplification, while sequencing of 12.870 bp through 32 loci in different genotypes revealed 85 SNP (one SNP every 151 bp). Seven SSRs were additionally tested revealing 5 polymorphic markers between donors and receivers. Polymorphisms were used to develop 35 PCR-based molecular markers suitable for introgressing of Pi genes into a set of the European rice germplasm. For this last purpose, allelic molecular marker variation was evaluated within a representative collection of about 95 rice genotypes. Polymorphic combinations allowing introgression of the broad spectrum resistance genes into a susceptible genetic background have been identified, thus confirming the potential of the identified markers for molecular-assisted breeding.     

Identification of Two Blast Resistance Genes in a Rice Variety, Digu

Blast, caused by Magnaporthe grisea is one of most serious diseases of rice worldwide. A Chinese local rice variety, Digu, with durable blast resistance, is one of the important resources for rice breeding for resistance to blast (M. grisea) in China. The objectives of the current study were to assess the identity of the resistance genes in Digu and to determine the chromosomal location by molecular marker tagging. Two susceptible varieties to blast, Lijiangxintuanheigu (LTH) and Jiangnanxiangnuo (JNXN), a number of different varieties, each containing one blast resistance gene, Pik^s, Pia, Pik, Pi‐b, Pi‐k^p, Pi‐ta², Pi‐ta, Pi‐z, Pi‐i, Pi‐k^m, Pi‐z^t, Pi‐t and Pi‐11, and the progeny populations from the crosses between Digu and each of these varieties were analysed with Chinese blast isolates. We found that the resistance of Digu to each of the two Chinese blast isolates, ZB13 and ZB15, were controlled by two single dominant genes, separately. The two genes are different from the known blast resistance genes and, therefore, designated as Pi‐d(t)1 and Pi‐d(t)2. By using bulked segregation method and molecular marker analysis in corresponding F₂ populations, Pi‐d(t)1 was located on chromosome 2 with a distance of 1.2 and 10.6 cM to restriction fragment length polymorphism (RFLP) markers G1314A and G45, respectively. And Pi‐d(t)2 was located on chromosome 6 with a distance of 3.2 and 3.4 cM to simple sequence repeat markers RM527 and RM3, respectively. We also developed a novel strategy of resistance gene analogue (RGA) assay with uneven polymerase chain reaction (PCR) to further tag the two genes and successfully identified two RGA markers, SPO01 and SPO03, which were co‐segregated toPi‐d(t)1 and Pi‐d(t)2, respectively, in their corresponding F₂ populations. These results provide essential information for further utilization of the Digu's blast resistance genes in rice disease resistance breeding and positional cloning of these genes.     

Phenotypic screening and molecular analysis of blast resistance in fragrant rice for marker assisted selection

Experiments were conducted to identify blast-resistant fragrant genotypes for the development of a durable blast-resistant rice variety during years 2012–2013. The results indicate that out of 140 test materials including 114 fragrant germplasms, 25 differential varieties (DVs) harbouring 23 blast-resistant genes, only 16 fragrant rice germplasms showed comparatively better performance against a virulent isolate of blast disease. The reaction pattern of single-spore isolate of Magnaporthe oryzae to differential varieties showed that Pish, Pi9, Pita-2 and Pita are the effective blast-resistant genes against the tested blast isolates in Bangladesh. The DNA markers profiles of selected 16 rice germplasms indicated that genotype Chinigura contained Pish, Pi9 and Pita genes; on the other hand, both BRRI dhan50 and Bawaibhog contained Pish and Pita genes in their genetic background. Genotypes Jirakatari, BR5, and Gopalbhog possessed Pish gene, while Uknimodhu, Deshikatari, Radhunipagol, Kalijira (3), Chinikanai each contained the Pita gene only. There are some materials that did not contain any target gene(s) in their genetic background, but proved resistant in pathogenicity tests. This information provided valuable genetic information for breeders to develop durable blast-resistant fragrant or aromatic rice varieties in Bangladesh.     

A set of near-isogenic lines of Indica-type rice variety CO 39 as differential varieties for blast resistance

Twenty-seven near-isogenic lines (NILs) with the genetic background of a blast-susceptible variety, CO 39, were developed by repeated backcrossing as a first set of a large number of differential varieties (DVs) with Indica-type genetic background. The NILs included 14 resistance genes—Pish, Pib, Piz-5, Piz-t, Pi5(t), Pik-s, Pik, Pik-h, Pik-m, Pik-p, Pi1, Pi7(t), Pita, and Pita-2—derived from 26 donor varieties. The reaction patterns of NILs against 20 standard isolates from the Philippines were similar to those of blast monogenic lines with the same resistance gene, except for those against two isolates that are avirulent to Pia in the genetic background of CO 39. A genome-wide DNA marker survey revealed that chromosome segments were introgressed in the regions where each resistance gene was previously mapped and most of the other chromosome regions in each NIL were CO 39 type. Segregation analysis of resistance and co-segregation analysis between resistance and DNA markers using F3 populations derived from the crosses between each NIL and the recurrent parent, CO 39, revealed a single-gene control of resistance and association between resistance and target introgressed segments. The morphological characters of each NIL were almost the same as those of the recurrent parent except for some lines, suggesting that these NILs can be used even under tropical conditions where Japonica-type DVs are not suitable for cropping. Thus, these NILs are useful not only as genetic tools for blast resistance study but also as sources of genes for breeding of Indica-type rice varieties.     

Development of PCR-based allele-specific and InDel marker sets for nine rice blast resistance genes

Blast resistance is one of the most important traits in rice breeding, and application of molecular markers for blast resistance breeding is likely to allow the rapid screening for the trait during early growth stages, without the need for inoculation of pathogen and phenotyping. Allele-specific PCR markers and insertion/deletion (InDel) markers, which genotype single-nucleotide polymorphisms and InDel polymorphisms, respectively, are useful tools for marker-assisted selections. We developed sets of allele-specific PCR and InDel markers for nine rice blast resistance genes—Piz, Piz-t, Pit, Pik, Pik-m, Pik-p, Pita, Pita-2, and Pib—which are commonly used in Japanese blast resistance rice breeding programs. For each resistance gene, we used the segregation information from thousands of progeny in several crosses or published gene locations to generate a marker that cosegregated with the gene and markers that closely flanked the gene on either side. The developed cosegregating markers uniquely discriminated among each of the lines with the individual resistance genes (except for Pita and Pita-2). Therefore, these markers will likely facilitate the development of multiline cultivars carrying one or a combination of these nine blast resistance genes. In addition, the systems we developed may be valuable tools in the quality control of seed production from blast-resistant multiline cultivars.     

Identification and fine mapping of <Emphasis Type="Italic">Pi33</Emphasis>, the rice resistance gene corresponding to the Magnaporthe grisea avirulence gene <Emphasis Type="Italic">ACE1</Emphasis>

Rice blast disease is a major constraint for rice breeding. Nevertheless, the genetic basis of resistance remains poorly understood for most rice varieties, and new resistance genes remain to be identified. We identified the resistance gene corresponding to the cloned avirulence gene ACE1 using pairs of isogenic strains of Magnaporthe grisea differing only by their ACE1 allele. This resistance gene was mapped on the short arm of rice chromosome 8 using progenies from the crosses IR64 (resistant) × Azucena (susceptible) and Azucena × Bala (resistant). The isogenic strains also permitted the detection of this resistance gene in several rice varieties, including the differential isogenic line C101LAC. Allelism tests permitted us to distinguish this gene from two other resistance genes [Pi11 and Pi-29(t)] that are present on the short arm of chromosome 8. Segregation analysis in F₂ populations was in agreement with the existence of a single dominant gene, designated as Pi33. Finally, Pi33 was finely mapped between two molecular markers of the rice genetic map that are separated by a distance of 1.6 cM. Detection of Pi33 in different semi-dwarf indica varieties indicated that this gene could originate from either one or a few varieties.Communicated by D.J. Mackill     

水稻抗稻瘟病基因Pi-d（t）^1、Pi-b、Pi-tα^2的聚合及分子标记选择

冈46B(G46B)是水稻生产应用中的一个农艺性状十分优良的保持系,其主要的缺陷是稻瘟病抗性较弱,通过对地谷,BL-1,Pi-4号等三个分别含抗病基因Pi-d(t)^1、Pi-b、Pi-tα^2的稻瘟病抗性材料与G46B聚合杂交,并利用抗病基因连锁的分子标记对杂交后代进行辅助选择,在聚合杂交的F2代及B1C1代群体中共获得了15株含Pi-d(t)^1、Pi-b、Pi-tα^2等三个抗稻瘟病基因的材料,其可能的基因型分别为：三基因杂合体Pi-d(t)^1pi-d(t)^1,Pi-bpi-b／Pi-tα^2 pi-tα^2 4株,双基因杂合体10株,其中Pi-d(t)^1Pi-d(t)^1／Pi-bpi-b／Pi-tα^2pi-tα^2 6株,Pi-d(t)^1pi-d(t)^1／Pi-bpi-b／Pi-tα^2Pi-tα^2 3株,Pi-d(t)^1pi-d(t)^1,Pi-bPi-6,Pi-tα^2 pi-tα^2 1株,双基因纯合体Pi-d(t)^1Pi-d(t)^1/Pi-bpi-b/Pi-tα^2Pi-tα^2仅1株,这一研究结果为进一步改良G46B的稻瘟病抗性奠定了基础,同时这一研究结果表明利用分子标记可快速、有效地实现多个抗病基因的聚合,大大提高水稻抗病育种的效率。     

Mapping of four major rice blast resistance genes from ’Lemont’ and ’Teqing’ and evaluation of their combinatorial effect for field resistance

A framework linkage map was developed using 284 F₁₀ recombinant inbred lines (RILs) from a ’Lemont’×’Teqing’ rice cultivar cross. Evaluation of a subset of 245 of these RILs with five races of the rice blast pathogen permitted RFLP mapping of three major resistance genes from Teqing and one major gene from Lemont. All mapped genes were found to confer resistance to at least two blast races, but none conferred resistance to all five races evaluated. RFLP mapping showed that the three resistance genes from Teqing, designated Pi-tq5, Pi-tq1 and Pi-tq6, were present on chromosomes 2, 6 and 12, respectively. The resistance gene from Lemont, Pi-lm2, was located on chromosome 11. Pi-tq1 is considered a new gene, based on its reaction to these five races and its unique map location, while the other three genes may be allelic with previously reported genes. Lines with different gene combinations were evaluated for disease reaction in field plots. Some gene combinations showed both direct effects and non-linear interaction. The fact that some of the lines without any of the four tagged genes exhibited useful levels of resistance in the field plots suggests the presence of additional genes or QTLs affecting the blast reaction segregating in this population. Received: 16 December 1999 / Accepted: 28 February 2000     

Molecular breeding: marker-assisted selection combined with biolistic transformation for blast and bacterial blight resistance in Indica rice (cv. CO39)

Based on blast pathogen population dynamics and lineage exclusion assays, we found that the major blast resistance genes Pi-1 and Piz-5 confer resistance against most Magnaporthe grisea lineages. Near-isogenic rice lines C101LAC and C101A51 carrying these two major genes for blast resistance in the background of a most blast-susceptible genotype were used for developing the pyramids. The closely linked RFLP marker RZ536 and NBS-LRR r10 marker for Pi-1 and a PCR-based SAP marker RG64 for Piz-5 were used to identify the genes in the parents and in marker-assisted breeding of the pyramided populations. To achieve multiple resistance against blast and blight in this cultivar, these blast-resistant pyramids were transformed with the cloned bacterial blight resistance gene Xa21 known to confer resistance to all races of Xanthomonas oryzae pv. oryzae (Xoo). Bioassays with six independent transformants showed that transgenic CO39 plants were resistant to both pathogens, M. grisea and Xoo. We report here the stacking of three major genes (Pi-1 + Piz-5 + Xa21) into rice using two different approaches of molecular breeding: marker-assisted selection (MAS) and genetic transformation.     

Functional markers based molecular characterization and cloning of resistance gene analogs encoding NBS-LRR disease resistance proteins in finger millet (<Emphasis Type="Italic">Eleusine coracana)</Emphasis>

Magnaporthe grisea, the blast fungus is one of the main pathological threats to finger millet crop worldwide. A systematic search for the blast resistance gene analogs was carried out, using functional molecular markers. Three-fourths of the recognition-dependent disease resistance genes (R-genes) identified in plants encodes nucleotide binding site (NBS) leucine-rich repeat (LRR) proteins. NBS-LRR homologs have only been isolated on a limited scale from Eleusine coracana. Genomic DNA sequences sharing homology with NBS region of resistance gene analogs were isolated and characterized from resistant genotypes of finger millet using PCR based approach with primers designed from conserved regions of NBS domain. Attempts were made to identify molecular markers linked to the resistance gene and to differentiate the resistant bulk from the susceptible bulk. A total of 9 NBS-LRR and 11 EST-SSR markers generated 75.6 and 73.5% polymorphism respectively amongst 73 finger millet genotypes. NBS-5, NBS-9, NBS-3 and EST-SSR-04 markers showed a clear polymorphism which differentiated resistant genotypes from susceptible genotypes. By comparing the banding pattern of different resistant and susceptible genotypes, five DNA amplifications of NBS and EST-SSR primers (NBS-05_504, NBS-09₇₁₁, NBS-07₆₈₈, NBS-03₅₀₉ and EST-SSR-04₂₄₁) were identified as markers for the blast resistance in resistant genotypes. Principal coordinate plot and UPGMA analysis formed similar groups of the genotypes and placed most of the resistant genotypes together showing a high level of genetic relatedness and the susceptible genotypes were placed in different groups on the basis of differential disease score. Our results provided a clue for the cloning of finger millet blast resistance gene analogs which not only facilitate the process of plant breeding but also molecular characterization of blast resistance gene analogs from Eleusine coracana.     

Rice (<i>Oryza sativa</i> L.) Breeding among Hassawi Landrace and Egyptian Genotypes for Stem Borer (<i>Chilo agamemnon</i> Bles.) Resistance and Related Quantitative Traits

Rice stem borer (Chilo agamemnon Bles.) is a primary insect pest of rice and is a major limiting factor to rice production. Breeding for insect-resistant crop varieties has been an economic way of integrated pest management (IPM) as it offers a viable and ecologically acceptable approach. This study was aimed to evaluate rice genotypes for their resistance against rice stem borer. Seven parental genotypes with twenty one F1 crosses were evaluated for genotypic variation in field experiments. Analysis of variance revealed significant differences for the studied traits in almost all crosses and parents. In addition, the mean squares of parents versus their crosses were signifi- cant for stem borer resistance and other associated traits. Moreover, both general combining ability (GCA) and specific combining ability (SCA) variances were highly significant for all characters studied in the F1 generation. Based on GCA, 4 genotypes (Sakha101, Gz6903-3-4-2-1, Gz9577-4-1-1 and Hassawi) exhibited highly significant negative values for stem borer resistance (–0.53, –1.06, –0.18 and –0.49, respectively) indicating they are the best combiners for stem borer resistance. Based on SCA analysis, nine cross combinations showed highly significant negative effects for stem borer resistance. Similarly, the cross Giza178 Hassawi was the best combination with significantly highest value for early maturity. In addition, seven crosses showed highly significant negative SCA for plant height trait. On the other hand, for panicle length, number of primary branches/panicle, panicle weight and 1000-grain weight, seven, four, eight and six crosses showed highly significant positive SCA, respectively. The result further revealed that the non-additive dominance genetic variance was higher than the additive variance for all evaluated traits indicating that non-additive genetic variances have a role in their inheritance. The broad-sense heritability estimates were high for all the studied traits. The stem borer resistance was significantly correlated with panicle weight and 1000-grain weight, which also showed a highly significant correlation with grain yield/plant. Thus these traits can be effectively employed in a breeding program to confer resistance against stem borer infestation in rice. It was further supported by biplot analysis, which clustered these potentially important traits into two quadrants showing their importance in any future breeding program to control stem borer infestation. This study has contributed valuable information for evaluation of genetic diversity in the local rice germplasm and its utilization in futuristic rice genetic improvement programs.     

Gene interactions and genetics of blast resistance and yield attributes in rice (Oryza sativa L.)

Blast disease caused by the pathogen Pyricularia oryzae is a serious threat to rice production. Six generations viz., P₁, P₂, F₁, F₂, B₁ and B₂ of a cross between blast susceptible high-yielding rice cultivar ADT 43 and resistant near isogenic line (NIL) CT13432-3R, carrying four blast resistance genes Pi1, Pi2, Pi33 and Pi54 in combination were used to study the nature and magnitude of gene action for disease resistance and yield attributes. The epistatic interaction model was found adequate to explain the gene action in most of the traits. The interaction was complementary for number of productive tillers, economic yield, lesion number, infected leaf area and potential disease incidence but duplicate epistasis was observed for the remaining traits. Among the genotypes tested under epiphytotic conditions, gene pyramided lines were highly resistant to blast compared to individuals with single genes indicating that the nonallelic genes have a complementary effect when present together. The information on genetics of various contributing traits of resistance will further aid plant breeders in choosing appropriate breeding strategy for blast resistance and yield enhancement in rice.     

Molecular markers linked to the blast resistance gene <Emphasis Type="Italic">Pi-z</Emphasis> in rice for use in marker-assisted selection

Rice blast, caused by the fungal pathogen Pyricularia grisea, is a serious disease affecting rice-growing regions around the world. Current methods for identification of blast-resistant germplasm and progeny typically utilize phenotypic screening. However, phenotypic screens are influenced by environmental conditions and the presence of one resistance gene can sometimes phenotypically mask other genes conferring resistance to the same blast race. Pi-z is a dominant gene located on the short arm of chromosome 6 that confers complete resistance to five races of blast. Using sequence data found in public databases and degenerate primer pairs based on the P-loop, nucleotide binding sites and kinase domain motifs of previously cloned resistance genes, we have developed PCR-based DNA markers that cosegregate with the gene. These markers are polymorphic in a wide range of germplasm, including the narrow crosses characteristic of applied rice-breeding programs. They can now be used as a low cost, high-throughput alternative to conventional phenotypic screening for direct detection of blast resistance genes, allowing rapid introgression of genes into susceptible varieties as well as the incorporation of multiple genes into individual lines for more-durable blast resistance.Electronic Supplementary Material Supplementary material is available in the online version of this article at Communicated by D. Mackill     

Combination Patterns of Major R Genes Determine the Level of Resistance to the M. oryzae in Rice (Oryza sativa L.)

Rice blast caused by Magnaporthe oryzae is the most devastating disease of rice and poses a serious threat to world food security. In this study, the distribution and effectiveness of 18 R genes in 277 accessions were investigated based on pathogenicity assays and molecular markers. The results showed that most of the accessions exhibited some degree of resistance (resistance frequency, RF >50%). Accordingly, most of the accessions were observed to harbor two or more R genes, and the number of R genes harbored in accessions was significantly positively correlated with RF. Some R genes were demonstrated to be specifically distributed in the genomes of rice sub-species, such as Pigm, Pi9, Pi5 and Pi1, which were only detected in indica-type accessions, and Pik and Piz, which were just harbored in japonica-type accessions. By analyzing the relationship between R genes and RF using a multiple stepwise regression model, the R genes Pid3, Pi5, Pi9, Pi54, Pigm and Pit were found to show the main effects against M. oryzae in indica-type accessions, while Pita, Pb1, Pik, Pizt and Pia were indicated to exhibit the main effects against M. oryzae in japonica-type accessions. Principal component analysis (PCA) and cluster analysis revealed that combination patterns of major R genes were the main factors determining the resistance of rice varieties to M. oryzae, such as ‘Pi9+Pi54’, ‘Pid3+Pigm’, ‘Pi5+Pid3+Pigm’, ‘Pi5+Pi54+Pid3+Pigm’, ‘Pi5+Pid3’ and ‘Pi5+Pit+Pid3’ in indica-type accessions and ‘Pik+Pib’, ‘Pik+Pita’, ‘Pik+Pb1’, ‘Pizt+Pia’ and ‘Pizt+Pita’ in japonica-type accessions, which were able to confer effective resistance against M. oryzae. The above results provide good theoretical support for the rational utilization of combinations of major R genes in developing rice cultivars with broad-spectrum resistance.     

Development and validation of co-dominant gene based markers for <Emphasis Type="Italic">Pi9</Emphasis>, a gene governing broad-spectrum resistance against blast disease in rice

Rice production and grain quality are severely affected by blast disease caused by the ascomycetous fungus Magnaporthe oryzae. Incorporation of genes that confer broad-spectrum resistance to blast has been a priority area in rice breeding programs. The blast resistance gene Pi9 sourced from Oryza minuta has shown broad spectrum and durable resistance to blast world-wide. In the present study co-dominant gene-based markers were developed for the precise marker-assisted tracking of Pi9 in breeding programs. The developed markers were validated across a diverse set of cultivars including basmati, indica and japonica varieties. Two markers, Pi9STS-1 and Pi9STS-2, effectively differentiated Pi9 donors from all the indicas and commercial basmati varieties tested. However, these markers were monomorphic between Pi-9 donors (IRBL9-W and Pusa 1637) and japonica type varieties. An additional gene-derived CAPS marker Pi91F_ 2R was developed to differentiate Pi9 donors from japonicas and traditional basmati lines. The co-dominant markers developed in the present study will be of immense utility to rice breeders for precise and speedy incorporation of Pi-9 into susceptible rice varieties through marker-assisted selection.     

Introgression of Pi2 and Pi5 Genes for Blast (Magnaporthe oryzae) Resistance in Rice and Field Evaluation of Introgression Lines for Resistance and Yield Traits

Blast caused by Magnaporthe oryzae is the most devastating disease causing significant loss in rice production. The destructive nature of the disease is mainly due to the genetic plasticity of M. oryzae which complicates the breeding strategies. Blast can be effectively managed by the deployment of R genes. In this study, broad‐spectrum blast resistance genes Pi2 and Pi5 were introgressed independently into popular but blast susceptible rice variety, Samba Mahsuri (BPT5204) by applying marker‐assisted backcross breeding approach. Tightly linked markers AP5930 for Pi2 and 40N23r for Pi5 gene were used in foreground selection. Background selection helped to identify the lines with maximum recovery of recurrent parent genome (RPG). The RPG recovery in Pi2 introgression lines was up to 90.17 and 91.46% in Pi5 lines. Homozygous introgression lines in BC₃F₄ generation carrying Pi2 and Pi5 gene were field evaluated for blast resistance, yield per se and yield‐related traits. The lines showed resistance to leaf and neck blast in multilocation field evaluation. Improved BPT5204 lines with improvement for blast resistance were on par with original BPT5204 in terms of grain yield and grain features.     

Genetic characterization and fine mapping of the blast resistance locus Pigm(t) tightly linked to Pi2 and Pi9 in a broad-spectrum resistant Chinese variety

The identification and utilization of broad-spectrum resistance genes have been proven the most effective and economical approach to control rice blast disease. To understand the molecular mechanism of broad-spectrum resistance to rice blast, we conducted genetic and fine mapping analysis of the blast resistance gene in a Chinese rice variety: Gumei 4 (GM4) identified with broad-spectrum resistance and used in rice breeding for blast resistance for more than 20 years. Genetic and mapping analysis indicated that blast resistance to nine isolates of different Chinese races in GM4 was controlled by the same dominant locus designated as Pigm(t) that was finely mapped to an approximately 70-kb interval between markers C5483 and C0428 on chromosome 6, which contains five candidate NBS--LRR disease resistance genes. The allelism test showed that Pigm(t) was either tightly linked or allelic to Pi2 and Pi9, two known blast resistance genes. Mapping information also indicated that another blast resistance gene Pi26(t) might also be located at the same region. Candidate genes were identified by sequence analysis of the Nipponbare and Pi9 locus and the corresponding region in GM4. Sequence divergence of candidate genes was observed between GM4 and model varieties Nipponbare and 9311, and Pi9. Our current study provides essential information and new genetic resource for the cloning of functional resistance gene(s) and for marker-assisted selection in rice breeding for broad-spectrum blast resistance.Yiwen Deng and Xudong Zhu contributed equally to this work.