Functional complementation of rice blast resistance gene Pi-k h (Pi54) conferring resistance to diverse strains of Magnaporthe oryzae

Abstract

Blast disease of rice, caused by Magnaporthe oryzae is an explosive disease that can spread rapidly in conducive conditions. R-gene mediated resistance offers an environmentally sustainable solution for management of this important disease of rice. We have earlier identified a unique R-gene of rice, on chromosome 11 of Oryza sativa ssp. indica cultivar Tetep. In this study we report functional validation of the Pi-k h (Pi54) gene using complementation assay. The blast resistance candidate gene Pi-k h (Pi54) was cloned into a plant transformation vector and the construct was used to transform a japonica cultivar of rice Taipei 309, which is susceptible to M. oryzae. Transgenic lines containing Pi-k h (Pi54) gene were found to confer high degree of resistance to diverse isolates of M. oryzae. The callose deposition was analyzed and compared between the transgenic and non-transgenic rice plants and widespread deposition was observed at the infection sites in plants showing incompatible interaction. Successful complementation of Pi-k h (Pi54) gene confirmed that the gene is responsible for resistance to M. oryzae in transgenic lines developed during this study. Expression analysis of the gene in resistant plants revealed that the gene is pathogen inducible in nature and is not expressed constitutively. Detection of callose deposition in resistant plants containing Pi-k h (Pi54) gene implicates its involvement in the initiation of defense response cascade.

A novel blast resistance gene, Pi54rh cloned from wild species of rice, Oryza rhizomatis confers broad spectrum resistance to Magnaporthe oryzae

“…Strategies aimed at breeding for durable blast resistance rice cultivars have focused on the possibility of stacking of multiple R-genes or transferring broad spectrum disease resistance genes in cultivated varieties ). In the present study, we used allele mining approach to clone a novel orthologue of durable rice blast resistance gene Pi54 (Rai et al 2011). In the present investigation, the expression of Pi54 orthologue was confirmed in the blast-resistant wild species of rice, O. rhizomatis using real-time PCR, postinoculation with the pathogen M. oryzae isolate PLP-1.…”

“…One of the recent approaches used for cloning new and better forms of disease resistance genes is allele mining for which the nucleotide sequence of the gene should be known. The dominant blast-resistant gene, Pi54 cloned from the indica rice line Tetep, associated with resistance to rice blast disease prevalent in different parts of India, has been mapped, cloned and characterized (Sharma et al 2005a, b;Rai et al 2011;Gupta et al 2012). Thus, allele mining for Pi54 orthologue from wild species of rice, O. rhizomatis can be a potential tool for finding alternate and better forms of the gene that can be used for rice blast management.…”

Rice Blast Management Through Host-Plant Resistance: Retrospect and Prospects

“…For instance, we have cloned blast resistance gene Pi54 from rice cultivar 'Tetep'. It shows broad spectrum resistance against several isolates prevalent in India [100]. We are also in the process of cloning of its counterpart avirulence gene, AvrPi54, from M. oryzae isolate RML-29 which is avirulent on Pi54 containing genotypes and the pathogen inducible promoter from rice plant itself which is responsive to a broad range of M. oryzae isolates and carry out early induction of downstream genes (unpublished findings).…”

“…Through complementation test, the Pi54 gene was found to confer resistance to four different isolates of the blast fungus in transgenic lines TP-Pi54-2 and TP-Pi54-15, among others. The gene was found to induce the synthesis of callose (b-1,3 glucan) in response to pathogen challenge, indicating its possible role in the initiation of a defense response cascade in the blast resistant plants [100]. The microarray and enzymatic analyses showed that Pi54 gene activates a cascade of defense response genes in a transgenic line up to T6 generation [35].…”

Identification of quantitative trait loci conferring blast resistance in Bodao, a japonica rice landrace

“…Most are dominant, except for the recessive gene pi21 (Jeung et al, 2007). Thus far, at least 20 resistance (R) genes have been cloned (Bryan et al, 2000;Liu et al, 2010;Okuyama et al, 2011;Rai et al, 2011;Yuan et al, 2011;Zhai et al, 2011;Hua et al, 2012), many of which have been organized into gene clusters, such as Pi2 loci (Pi2/ Piz-t/Pi9) (Deng et al, 2006;Qu et al, 2006;Zhou et al, 2006) and Pik loci (Pik/Pik-m/ Pik-p/Pi1) (Ashikawa et al, 2008;Yuan et al, 2011;Zhai et al, 2011;Hua et al, 2012). Among the 20 cloned R genes, 18 genes encode nucleotide binding sites and leucine-rich repeat proteins, except for Pi-d2, which encodes a B-lectin receptor kinase (Chen et al, 2006), and pi21, which encodes a proline-rich protein that contains a putative heavy metalbinding domain and protein-protein interaction motifs (Fukuoka et al, 2009).…”