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Dual gene expression cassette is superior than single gene cassette for enhancing sheath blight tolerance in transgenic rice.

ABSTRACT: Sheath blight, caused by the necrotrophic fungal pathogen Rhizoctonia solani, is a serious and destructive disease of the rice. In order to improve sheath blight resistance, we developed three different kinds of transgenic rice lines. The first transgenic line overexpresses the rice chitinase gene (OsCHI11); the second contains the Arabidopsis NPR1 (AtNPR1) gene and, the third has pyramided constructs with both the genes (OsCHI11 and AtNPR1). This is a comparative study between the single-gene transgenic lines and the double gene transgenic in terms of their ability to activate the plant defense system. Rice plants of each individual construct were screened via PCR, Southern hybridization, activity assays, and expression analysis. The best transgenic lines of each construct were chosen for comparative study. The fold change in qRT-PCR and activity assays revealed that the pyramided transgenic rice plants show a significant upregulation of defense-related genes, PR genes, and antioxidant marker genes as compared to the single transgene. Simultaneous co-expression of both the genes was found to be more efficient in tolerating oxidative stress. In R. solani (RS) toxin assay, mycelial agar disc bioassay, and in vivo plant bioassay, pyramided transgenic plant lines were more competent at restricting the pathogen development and enhancing sheath blight tolerance as compared to single gene transformants.

SUBMITTER: Karmakar S

PROVIDER: S-EPMC5554252 | biostudies-literature | 2017 Aug

REPOSITORIES: biostudies-literature

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Dual gene expression cassette is superior than single gene cassette for enhancing sheath blight tolerance in transgenic rice.

Karmakar Subhasis S Molla Kutubuddin A KA Das Kaushik K Sarkar Sailendra Nath SN Datta Swapan K SK Datta Karabi K

Scientific reports 20170811 1

Sheath blight, caused by the necrotrophic fungal pathogen Rhizoctonia solani, is a serious and destructive disease of the rice. In order to improve sheath blight resistance, we developed three different kinds of transgenic rice lines. The first transgenic line overexpresses the rice chitinase gene (OsCHI11); the second contains the Arabidopsis NPR1 (AtNPR1) gene and, the third has pyramided constructs with both the genes (OsCHI11 and AtNPR1). This is a comparative study between the single-gene t ...[more]

PMID: 28801565

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Project description:Bacterial blight, blast, and sheath blight are the commonest diseases causing substantial yield loss in rice around the world. Stacking of broad-spectrum resistance genes/QTLs into popular cultivars is becoming a major objective of any disease resistance breeding program. The varieties ASD 16 and ADT 43 are the two popular, high yielding, and widely grown rice cultivars of South India, which are susceptible to bacterial blight (BB), blast, and sheath blight diseases. The present study was carried out to improve the cultivars (ASD 16 and ADT 43) through introgression of bacterial blight (xa5, xa13, and Xa21), blast (Pi54), and sheath blight (qSBR7-1, qSBR11-1, and qSBR11-2) resistance genes/QTLs by MABB (marker-assisted backcross breeding). IRBB60 (xa5, xa13, and Xa21) and Tetep (Pi54; qSBR7-1, qSBR11-1, and qSBR11-2) were used as donors to introgress BB, blast, and sheath blight resistance into the recurrent parents (ASD 16 and ADT 43). Homozygous (BC3F3 generation), three-gene bacterial blight pyramided (xa5 + xa13 + Xa21) lines were developed, and these lines were crossed with Tetep to combine blast (Pi54) and sheath blight (qSBR7-1, qSBR11-1, and qSBR11-2) resistance. In BC3F3 generation, the improved pyramided lines carrying a total of seven genes/QTLs (xa5 + xa13 + Xa21 + Pi54 + qSBR7-1 + qSBR11-1 + qSBR11-2) were selected through molecular and phenotypic assay, and these were evaluated for resistance against bacterial blight, blast, and sheath blight pathogens under greenhouse conditions. We have selected nine lines in ASD 16 background and 15 lines in ADT 43 background, exhibiting a high degree of resistance to BB, blast, and sheath blight diseases and also possessing phenotypes of recurrent parents. The improved pyramided lines are expected to be used as improved varieties or used as a potential donor in breeding programs. The present study successfully introgressed Pi54, and qSBR QTLs (qSBR7-1, qSBR11-1, and qSBR11-2) from Tetep and major effective BB-resistant genes (xa5, xa13, and Xa21) from IRBB60 into the commercial varieties for durable resistance to multiple diseases.

Project description:Potential biological control agents for two major rice diseases, sheath blight and bacterial panicle blight, were isolated from rice plants in this study. Rice-associated bacteria (RABs) isolated from rice plants grown in the field were tested for their antagonistic activities against the rice pathogens, Rhizoctonia solani and Burkholderia glumae, which cause sheath blight and bacterial panicle blight, respectively. Twenty-nine RABs were initially screened based on their antagonistic activities against both R. solani and B. glumae. In follow-up retests, 26 RABs of the 29 RABs were confirmed to have antimicrobial activities, but the rest three RABs did not reproduce any observable antagonistic activity against R. solani or B. glumae. According to16S rDNA sequence identity, 12 of the 26 antagonistic RABs were closest to Bacillus amyloliquefaciens, while seven RABs were to B. methylotrophicus and B, subtilis, respectively. The 16S rDNA sequences of the three non-antagonistic RABs were closest to Lysinibacillus sphaericus (RAB1 and RAB12) and Lysinibacillus macroides (RAB5). The five selected RABs showing highest antimicrobial activities (RAB6, RAB9, RAB16, RAB17S, and RAB18) were closest to B. amyloliquefaciens in DNA sequence of 16S rDNA and gyrB, but to B. subtilis in that of recA. These RABs were observed to inhibit the sclerotial germination of R. solani on potato dextrose agar and the lesion development on detached rice leaves by artificial inoculation of R. solani. These antagonistic RABs also significantly suppressed the disease development of sheath blight and bacterial panicle blight in a field condition, suggesting that they can be potential biological control agents for these rice diseases. However, these antagonistic RABs showed diminished disease suppression activities in the repeated field trial conducted in the following year probably due to their reduced antagonistic activities to the pathogens during the long-term storage in -70C, suggesting that development of proper storage methods to maintain antagonistic activity is as crucial as identification of new biological control agents.

Dataset Information

Dual gene expression cassette is superior than single gene cassette for enhancing sheath blight tolerance in transgenic rice.

Publications

Dual gene expression cassette is superior than single gene cassette for enhancing sheath blight tolerance in transgenic rice.

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