The over-expression of a glycine-rich RNA-binding protein in Nicotiana tabacum and its roles in plant stress tolerance

Glycine-rich RNA-binding proteins (GR-RBPs) have been explored in different plant species for their role in development and environmental stress responses. NtGR-RBP1, a tobacco glycine-rich RNA-binding protein has recently been suggested as RNA chaperone with reference to protective role in bacteria upon exposure to various stresses. In the present study, we aimed to test the roles of NtGR-RBP1 in plants. The cDNA sequence of NtGR-RBP1 was amplified and cloned into entry vector using pENTR/D-TOPO cloning kit followed by sub-cloning into a plant GATEWAY (R) expression vector resulting in construction of pH7WG2-NtGR-RBP1. NtGR-RBP1 recombinants were confirmed through colony PCR and sequencing. Further, pH7WG2-NtGR-RBP1 was cloned into Agrobacterium tumefaciens AGL1 competent cells and confirmed by PCR after screening on streptomycin and hygromycin. The culture was subsequently used for the transformation of Nicotiana tabacum cv samsun by Agrobacterium-mediated plant transformation using the leaf disc method. Explants were screened using hygromycin as selection marker and later confirmed through PCR. Seed germination assay showed that NtGR-RBP1 had significant role in response to cold, heat and drought stress, while salt stress has negative effect on germination. Transformed plants will be used further for in depth functional characterization of NtGR-RBP1.