Pectin induced transcriptome of a Rhizoctonia solani strain causing sheath blight disease in rice reveals insights on key genes and RNAi machinery for development of pathogen derived resistance

Key message RNAi mediated silencing of pectin degrading enzyme of R. solani gives a high level of resistance against sheath blight disease of rice. Abstract Rice sheath blight disease caused by Rhizoctonia solani Kuhn (telemorph; Thanatephorus cucumeris) is one of the most devastating fungal diseases which cause severe loss to rice grain production. In the absence of resistant cultivars, the disease is currently managed through fungicides which add to environmental pollution. To explore the potential of utilizing RNA interference (RNAi)-mediated resistance against sheath blight disease, we identified genes encoding proteins and enzymes involved in the RNAi pathway in this fungal pathogen. The RNAi target genes were deciphered by RNAseq analysis of a highly virulent strain of the R. solani grown in pectin medium. Additionally, pectin metabolism associated genes of R. solani were analyzed through transcriptome sequencing of infected rice tissues obtained from six diverse rice cultivars. One of the key candidate gene AG1IA_04727 encoding polygalacturonase (PG), which was observed to be significantly upregulated during infection, was targeted through RNAi to develop disease resistance. Stable expression of PG-RNAi construct in rice showed efficient silencing of AG1IA_04727 and suppression of sheath blight disease. This study highlights important information about the existence of RNAi machinery and key genes of R. solani which can be targeted through RNAi to develop pathogen-derived resistance, thus opening an alternative strategy for developing sheath blight-resistant rice cultivars

Development and evaluation of introgression lines with yield enhancing genes of the Indian mega-variety of rice, MTU1010

MTU 1010 is an early maturing and high-yielding mega rice variety widely grown in an area of 3 Mha. It is characterised by limited grain number and panicle branching. To improve the grain number in MTU 1010, an IRRI breeding line, IR121055-2-10-5 was utilized as donor to transfer yield-enhancing genes Gn1a and OsSPL14 (associated with increased grain number and better panicle branching, respectively) into MTU1010 by Marker-Assisted Backcross Breeding (MABB). At each backcross generation, foreground selection was carried out with Gn1a and OsSPL14- specific molecular markers, whilst background selection was done with a set of SSR markers polymorphic between the IR121055-2-10-5 and MTU1010. With the use of a gene-specific marker, homozygous BC2 F2 plants carrying the yield-enhancing gene were identified and advanced through pedigree-method of selection till BC2 F6 and best performing ten lines were selected and evaluated in replicated station trials for yield contributing traits, where grain number and brancing per panicle exhibited high significant and positive correlation with single plant yield. Three promising lines namely RP6353-5-8-13-24, RP6353-26-13-39-5 and RP6353-32-12-8-16 with higher grain number and yield than MTU1010 were identified and nominated for evaluation in Initial Varietal Trial-Aerobic (IVT-Aerobic) of All India Crop Improvement Programme on Rice (AICRP), of which RP6353-26-13-39-5 (IET28674), was promoted for further testing

Phenotypic and molecular confirmation of durable adult plant leaf rust resistance (APR) genes Lr34+, Lr46+ and Lr67+ linked to leaf tip necrosis (LTN) in select registered Indian wheat (T. aestivum) genetic stocks

Nearly twenty thousand wheat lines were phenotyped for the presence of leaf tip necrosis (LTN), a phenotypic trait linked to adult plant leaf rust resistance (APR) genes, viz. Lr34, Lr46 and Lr67 having pleiotropic association with multiple disease resistance genes. Thirty-six lines showed varied expression of LTN and moderate level of leaf rust severity at adult plant stage with slow rusting (disease progress at a retarded rate). Seedling resistance test (SRT) revealed susceptible and mixed infection types, a characteristic of adult plant resistance (APR) genes. Further molecular confirmation for the presence of these genes using available microsatellite markers revealed that of the 36 lines, five lines carried Lr46+ alone and five other lines carried Lr67+ alone. Seven lines carried the combination of Lr34+ and Lr67+ while six lines confirmed to carry the combination of Lr46+ and Lr67+. Remarkably three lines carried all the three APR genes, viz. Lr34+, Lr46+ and Lr67+. All these stocks can be a source of APR multiple disease resistance genes. Ten lines were not confirmed to carry any of the genes but still had LTN and SRT results showing an infection type typical of APR genes and these can be the source of identifying newer APR genes. The resistance based on minor APR genes when combined with a few additional minor genes in the background of high yielding cultivars is expected to have high level of race non-specific resistance and to be durable

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Not AvailableNot RNAi mediated silencing of pectin degrading enzyme of R. solani gives a high level of resistance against sheath blight disease of rice. Rice sheath blight disease caused by Rhizoctonia solani Kuhn (telemorph; Thanatephorus cucumeris) is one of the most devastating fungal diseases which cause severe loss to rice grain production. In the absence of resistant cultivars, the disease is currently managed through fungicides which add to environmental pollution. To explore the potential of utilizing RNA interference (RNAi)-mediated resistance against sheath blight disease, we identified genes encoding proteins and enzymes involved in the RNAi pathway in this fungal pathogen. The RNAi target genes were deciphered by RNAseq analysis of a highly virulent strain of the R. solani grown in pectin medium. Additionally, pectin metabolism associated genes of R. solani were analyzed through transcriptome sequencing of infected rice tissues obtained from six diverse rice cultivars. One of the key candidate gene AG1IA_04727 encoding polygalacturonase (PG), which was observed to be significantly upregulated during infection, was targeted through RNAi to develop disease resistance. Stable expression of PG-RNAi construct in rice showed efficient silencing of AG1IA_04727 and suppression of sheath blight disease. This study highlights important information about the existence of RNAi machinery and key genes of R. solani which can be targeted through RNAi to develop pathogen-derived resistance, thus opening an alternative strategy for developing sheath blight-resistant rice cultivars.Not Availabl

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Not AvailablePhosphorus (P) is an essential macronutrient to all crops including rice and it plays a key role in various plant activities and development. Low availability of P in the soils negatively, influences rice crop growth and causes significant yield loss. In the present study, we characterized a set of 56 germplasm lines for their tolerance to low soil P by screening them at low soil P and optimum soil P levels along with low soil P tolerant and sensitive check varieties. These lines were genotyped for the presence/absence of tolerant allele with respect to the major low soil P tolerance QTL, Pup1, using a set of locus specific PCR-based markers, viz., K46-1, K46-2, K52 and K46CG-1. High genetic variability was observed for various traits associated with low soil P tolerance. The yield parameters from normal and low soil P conditions were used to calculate stress tolerance indices and classify the genotypes according to their tolerance level. Out of the total germplasm lines screened, 15 lines were found to be tolerant to low soil P condition based on the yield reduction in comparison to the tolerant check, but most of them harbored the complete or partial Pup1 locus. Interestingly, two tolerant germplasm lines, IC216831 and IC216903 were observed to be completely devoid of Pup1 and hence they can be explored for new loci underlying low soil P tolerance.Not Availabl

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Not AvailableMTU1010 is a high-yielding mega-variety of rice grown extensively in India. However, it does not perform well in soils with low phosphorus (P) levels. With an objective to improve MTU 1010 for tolerance to low soil P, we have transferred Pup1, a major quantitative trait locus (QTL) associated with tolerance from another mega-variety, Swarna, through marker-assisted back cross breeding (MABB). Foreground selection of the F1 and backcross plants was performed with the codominant, closely linked CAPS marker, K20-2, while two flanking markers RM28011 and RM28157 were utilized for recombinant selection. At each backcross generation, positive plants were also analyzed with a set of 85 parental polymorphic SSR markers to identify theQTL-positive plants possessing maximum introgression of MTU 1010 genome. At BC2F1, the best backcross plant was selfe dtogenerate BC2F2s. Among them, the plants homozygous for Pup1 (n = 22) were reconfirmed using the functional marker for Pup1, viz., K46-1, and they were advanced through pedigree method of selection until BC2F6 generation. A total of five elite BC2F6 lines, possessing Pup1 and phenotypically similar to MTU 1010, were screened in the low soil P plot and normal plot (with optimum soil P levels) during wet season, 2016. All the selected lines showed better performance under low P soil with more number of productive tillers,better root system architecture,and significantly higher yield (>390%) as compared to MTU 1010. Further, under normal soil, the lines were observed to be similar to or better than MTU 1010 for most of the agro-morphological traits and yield. This study represents the successful application of marker assisted selection for improvement of tolerance to low soil P in a high-yielding Indian rice variety.The authors received financial support provided by the Department of Biotechnology (DBT), Government of India, for execution of the research study through the Grant # BT/PR4665/AGII/106/854/2012 dated: 19/02/2013

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Not AvailableImproved Samba Mahsuri (ISM) is a popular, high-yielding, bacterial blight resistant rice variety possessing medium-slender grain type. As ISM is highly susceptible to blast disease of rice, through the present study we have transferred two major blast resistance genes, Pi2 and Pi54 into the elite variety by marker-assisted backcross breeding. The two blast resistance genes were transferred to ISM through sets of backcrosses. In every backcross generation, PCR-based markers, specific for the blast resistance genes (Pi2 and Pi54) and bacterial blight resistance genes (Xa21, xa13 and xa5) were utilized for foreground selection, while a set of 144 parental polymorphic SSR markers were used for background selection and backcrossing was carried out until BC2 generation. A solitary BC2F1 plant possessing Pi2 or Pi54 along with Xa21, xa13 and xa5 and > 90% recovery of ISM genome was selected from the two sets of backcrosses were crossed and the intercross F1s (ICF1s) thus obtained were selfed to generate ICF2s. Homozygous ICF2 plants carrying all the five resistance genes were identified through markers and advanced through selfing till ICF5 generation by adopting pedigree method of selection. Three best lines at ICF5, possessing excellent resistance against bacterial blight and blast and closely resembling or superior to ISM in terms of grain quality: yield and agro-morphological traits have been identified and advanced for multi-location trials.ICAR-Indian Agricultural Research Institute with award Number: (F.No.F.3/CRPMB/Gen/2015-16/1714)Department of Science and Technology (DST), Government of India for the INSPIRE fellowshi