Molecular analysis of stress response dynamics in rice against Rice tungro viruses and assessment of RNAi mediated viral resistance in transgenic and backcrossed rice varieties

Abstract

Rice tungro is the most damaging viral disease of rice caused by two viruses: Rice Tungro Bacilliform Virus and Rice Tungro Spherical Virus through the Green Leafhopper (GLH) (Nephotettix virescens) vector. The transcriptome study of tungro infected rice revealed about 1000 differentially regulated genes, mapped on several pathways such as those of jasmonic acid, secondary metabolites and ROS metabolism etc. Out of the 17 targets of the differentially regulated miRNA obtained by small RNA sequencing of tungro infected plants, 9 targets expression exactly correlated with the expression of their corresponding miRNA.The expression analysis of RNAi components in rice upon tungro infection by RT-PCR revealed four DCLs, five AGOs and three RDRs to be differentially regulated upon tungro infection their levels exactly correlated with level of virus in different plant parts; silencing of these genes is achieved through VIGS-mediated silencing. The expression analysis of the host cell wall-related genes and components revealed that the tungro infection causes suppression of these genes and study of host ROS machinery upon tungro infection indicated how tungro infection causes an imbalance in the host ROS homeostasis resulting into degradation symptoms. The resistance assay of the transgenic and backcrossed rice varieties having a double-stranded RNA construct against RTBV revealed three lines of ASD 16, two lines each of BPT 5204 and CR 1009 and one line of Shatabdi to be RTBV resistant. Dual RNAi strategy in transgenic plants against tungro virus complex was used which resulted into resistance against both RTBV and RTSV.