Research Projects

Crop improvement has been largely relying on the natural genetic variation. However, habitat loss and overuse of existing germplasm have narrowed this genetic base, limiting the potential of conventional breeding approaches. There is a growing need for precise, programmable tools to manipulate gene expression in both model and crop plants. M1GS ribozyme, an engineered bacterial RNase P, is a versatile and a simple RNA-cleaving tool involving a single catalytic RNA (~0.5kb), has been used for targeted downregulation of transcripts in a sequence specific manner in bacteria and mammalian cells. However, it remains unexplored in plant systems. Additionally, epigenetic variation especially through histone deacetylase (HDAC) regulation has shown promising results as a new modality for crop improvement, as suggested by recent literature (Xu et al., 2022, Nguyen et al., 2018). In rice, the histone deacetylase gene OsHDA705 is implicated in abiotic stress responses, and its downregulation could potentially enhance stress tolerance. By integrating M1GS technology with functional epigenetic knockdown, this project aims to develop novel tool for reverse genetics and improve crop tolerance to environmental stresses. For this purpose, the scientific objectives of this project are centered around establishing and applying M1GS ribozyme technology in plants. Preliminary data we have obtained suggest that M1GS can function in plants but thorough studies are required to obtain highly efficient version of M1GS and appropriate vector system. Therefore, different M1GS variants and different vector systems will be tested for their expression and RNA-cleaving efficiency by targeting the PDS gene in Nicotiana benthamiana (a model dicot plant species). Next, the study aims to demonstrate targeted downregulation of OsHDA705, a histone deacetylase gene, in a popular Indian rice variety Improved Samba Mahsuri (ISM) using M1GS constructs. Finally, the resulting rice lines with downregulated OsHDA705 expression will be evaluated for their tolerance to abiotic stresses such as drought and salinity. We hypothesize that bacterial-derived M1GS ribozymes can function effectively in plant systems to cleave target RNAs in a sequence-specific manner. Our recent data in mammalian cells suggest that M1GS could be more efficient than RNAi and CRISPR/Cas13(unpublished data). We also hypothesize that downregulation of OsHDA705 would lead to enhanced abiotic stress tolerance in rice, presumably by derepressing stress responsive genes. Based on the phenotypic data RNAseq analysis will be carried out in these lines to identify differentially regulated genes in Oshda705 knockdown lines. In summary, the project has the potential to establish M1GS ribozymes as a novel RNA-cleaving tool for plants, providing an alternative to RNAi and CRISPR-based approaches. The findings could open new avenues for functional genomics, reverse genetics, and epigenetic crop improvement strategies.