Advances in breeding, biotechnology, and nanotechnological approaches to combat sheath blight disease in rice.

Sheath blight, caused by the fungus Rhizoctonia solani, is a major problem that significantly impacts rice production and can lead to substantial yield losses. The disease has become increasingly problematic in recent years due to the widespread use of high-yielding semi-dwarf rice cultivars, dense planting, and heavy application of nitrogenous fertilizers. The disease has become more challenging to manage due to its diverse host range and the lack of resistant cultivars. Despite utilizing traditional methods, the problem persists without a satisfactory solution. Therefore, modern approaches, including advanced breeding, transgenic methods, genome editing using CRISPR/Cas9 technology, and nanotechnological interventions, are being explored to develop rice plants resistant to sheath blight disease. This review primarily focuses on these recent advancements in combating the sheath blight disease.

Strategies for Efficient RNAi-Based Gene Silencing of Viral Genes for Disease Resistance in Plants.

RNA interference (RNAi) is an evolutionarily conserved gene silencing mechanism in eukaryotes including fungi, plants, and animals. In plants, gene silencing regulates gene expression, provides genome stability, and protect against invading viruses. During plant virus interaction, viral genome derived siRNAs (vsiRNA) are produced to mediate gene silencing of viral genes to prevent virus multiplication. After the discovery of RNAi phenomenon in eukaryotes, it is used as a powerful tool to engineer plant viral disease resistance against both RNA and DNA viruses. Despite several successful reports on employing RNA silencing methods to engineer plant for viral disease resistance, only a few of them have reached the commercial stage owing to lack of complete protection against the intended virus. Based on the knowledge accumulated over the years on genetic engineering for viral disease resistance, there is scope for effective viral disease control through careful design of RNAi gene construct. The selection of target viral gene(s) for developing the hairpin RNAi (hp-RNAi) construct is very critical for effective protection against the viral disease. Different approaches and bioinformatics tools which can be employed for effective target selection are discussed. The selection of suitable target regions for RNAi vector construction can help to achieve a high level of transgenic virus resistance.

Combination of driselase and lysing enzyme in one molar potassium chloride is effective for the production of protoplasts from germinated conidia of Fusarium verticillioides.

Various cell wall degrading enzymes and the protoplasting media were evaluated for the production of protoplast in Fusarium verticillioides. Among the various enzymes tested, driselase at 12.5 mg/ml in 1 M KCl protoplasting medium produced the maximum number of protoplast. Next to driselase, lysing enzyme at 10 mg/ml in 1.2 M MgSO4 protoplasting medium was found to be the second best enzyme for the production of protoplast. More interestingly, the combined use of driselase @ 12.5 mg/ml and lysing enzyme @ 10 mg/ml in 1 M KCl exhibited the additive effect on protoplast formation. Germinated conidia of F. verticillioides are the most susceptible fungal material for protoplast production. The use of sucrose at 1.2 M in the regeneration medium supported the maximum regeneration of protoplast. From the present study, we recommend driselase (12.5 mg/ml) and lysing enzyme (10 mg/ml) in 1 M KCl protoplasting medium and germinated conidia of F. verticillioides for the maximum production of protoplasts and 1.2 M sucrose is the best osmoticum for the regeneration of protoplasts.

Development of Agrobacterium-mediated transformation of highly valued hill banana cultivar Virupakshi (AAB) for resistance to BBTV disease.

One of the most severe viral diseases of hill banana is caused by banana bunchy top virus (BBTV), a nanovirus transmitted by the aphid Pentalonia nigronervosa. In this study, we reported the Agrobacterium-mediated transformation on a highly valued hill banana cultivar Virupakshi (AAB) for resistance to BBTV disease. The target of the RNA interference (RNAi) is the rep gene, encoded by the BBTV-DNA1. In order to develop RNAi construct targeting the BBTV rep gene, the full-length rep gene of 870 bp was polymerase chain reaction amplified from BBTV infected hill banana sample DNA, cloned and confirmed by DNA sequencing. The partial rep gene fragment was cloned in sense and anti sense orientation in the RNAi intermediate vector, pSTARLING-A. After cloning in pSTARLING-A, the cloned RNAi gene cassette was released by NotI enzyme digestion and cloned into the NotI site of binary vector, pART27. Two different explants, embryogenic cells and embryogenic cell suspension derived microcalli were used for co-cultivation. Selection was done in presence of 100 mg/L kanamycin. In total, 143 putative transgenic hill banana lines were generated and established in green house condition. The presence of the transgenes was confirmed in the selected putative transgenic hill banana lines by PCR and reverse transcription PCR analyses. Transgenic hill banana plants expressing RNAi-BBTV rep were obtained and shown to resist infection by BBTV. The transformed plants are symptomless, and the replication of challenge BBTV almost completely suppressed. Hence, the RNAi mediating resistances were shown to be effective management of BBTV in hill banana.

The mediator complex subunit PFT1 is a key regulator of jasmonate-dependent defense in Arabidopsis.

Jasmonate signaling plays an important role in both plant defense and development. Here, we have identified a subunit of the Mediator complex as a regulator of the jasmonate signaling pathway in Arabidopsis thaliana. The Mediator complex is a conserved multiprotein complex that acts as a universal adaptor between transcription factors and the RNA polymerase II transcriptional machinery. We report that the PHYTOCHROME AND FLOWERING TIME1 (PFT1) gene, which encodes the MEDIATOR25 subunit of Mediator, is required for jasmonate-dependent defense gene expression and resistance to leaf-infecting necrotrophic fungal pathogens. Conversely, PFT1 appears to confer susceptibility to Fusarium oxysporum, a root-infecting hemibiotrophic fungal pathogen known to hijack jasmonate responses for disease development. Consistent with this, jasmonate gene expression was suppressed in the pft1 mutant during infection with F. oxysporum. In addition, a wheat (Triticum aestivum) homolog of PFT1 complemented the defense and the developmental phenotypes of the pft1 mutant, suggesting that the jasmonate signaling functions of PFT1 may be conserved in higher plants. Overall, our results identify an important control point in the regulation of the jasmonate signaling pathway within the transcriptional machinery.