Hairpin RNA Targeting Multiple Viral Genes Confers Strong Resistance to Rice Black-Streaked Dwarf Virus.

Fangquan Wang,Tong Zhou,Jinyan Zhu,Qing Liu,Fangjun Fan,Qian-Hao Zhu,Jie Yang,Weigong Zhong,Jun Wang,Yijun Zhou,Wenqi Li,Ying Lan,Ming-Bo Wang

doi:10.3390/ijms17050705

Abstract

Rice black-streaked dwarf virus (RBSDV) belongs to the genus Fijivirus in the family of Reoviridae and causes severe yield loss in rice-producing areas in Asia. RNA silencing, as a natural defence mechanism against plant viruses, has been successfully exploited for engineering virus resistance in plants, including rice. In this study, we generated transgenic rice lines harbouring a hairpin RNA (hpRNA) construct targeting four RBSDV genes, S1, S2, S6 and S10, encoding the RNA-dependent RNA polymerase, the putative core protein, the RNA silencing suppressor and the outer capsid protein, respectively. Both field nursery and artificial inoculation assays of three generations of the transgenic lines showed that they had strong resistance to RBSDV infection. The RBSDV resistance in the segregating transgenic populations correlated perfectly with the presence of the hpRNA transgene. Furthermore, the hpRNA transgene was expressed in the highly resistant transgenic lines, giving rise to abundant levels of 21–24 nt small interfering RNA (siRNA). By small RNA deep sequencing, the RBSDV-resistant transgenic lines detected siRNAs from all four viral gene sequences in the hpRNA transgene, indicating that the whole chimeric fusion sequence can be efficiently processed by Dicer into siRNAs. Taken together, our results suggest that long hpRNA targeting multiple viral genes can be used to generate stable and durable virus resistance in rice, as well as other plant species.

Highlights

Rice is one of the major cereal crops in the world and is important for global food security
The hairpin RNA (hpRNA) transgenic lines were assayed for rice black-streaked dwarf virus (RBSDV) resistance using T1, T2 and T3 populations
A greater difference was observed in 2014: the wild-type plants displayed a ~41.44% disease incidence rate, whereas the transgenic lines showed only a ~0%–14% disease rate (Table S3). These results suggested that the transgenic rice lines have enhanced RBSDV resistance

Summary

Introduction

Rice is one of the major cereal crops in the world and is important for global food security. Rice black-streaked dwarf virus disease is a destructive viral disease, caused by rice black-streaked dwarf virus (RBSDV), which mainly infects rice and maize plants in East Asian countries [1,2,3] This virus disease caused serious yield losses of rice and maize in Japan during the 1960s [4]. A disease epidemic by RBSDV depends on the abundance of viruliferous hosts and the number of SBPH carrying RBSDV. These characteristics of RBSDV transmission lead to the difficulty in screening for RBSDV-resistant rice germplasms and evaluating the resistance level of rice materials. Assay results from the artificial inoculation method are generally reliable, it is time-consuming and laborious for large-scale screening and evaluation of rice collections [6,7]

Methods

Results

Conclusion