Generating Novel Male Sterile Tomatoes by Editing Respiratory Burst Oxidase Homolog Genes
Xiaojuan Dai, Huanan Han, Wei Huang, Lianghui Zhao, Ming-Lei Song, Xuesong Cao, Chenglan Liu, Xiaomu Niu, Zhaobo Lang, Changle Ma, Hongtao Xie
Abstract
Hybrid breeding of tomatoes ( Solanum lycopersicum ), an important vegetable crop, is an effective way to improve yield and enhance disease and stress resistance. However, the efficiency of tomato hybridization is hindered by self-fertilization, which can be overcome using male sterile lines. It has been reported that reactive oxygen species (ROS) act as a key regulator for anther development, mediated by RBOH ( Respiratory Burst Oxidase Homolog ) genes. Here, two tomato anther-expressed genes, LeRBOH (Solyc01g099620) and LeRBOHE (Solyc07g042460), were selected to cultivate novel tomato male sterile strains. By using a CRISPR/Cas9 system with a two-sgRNA module, the lerboh , lerbohe , and lerboh lerbohe mutant lines were generated, among which the lerbohe and lerboh lerbohe mutants displayed complete male sterility but could accept wild-type pollens and produce fruits normally. Further analysis uncovered significantly decreased ROS levels and abnormal programmed cell death in lerboh lerbohe anthers, indicating a key role of ROS metabolism in tomato pollen development. Taken together, our work demonstrates a successful application of gene editing via CRISPR/Cas9 in generating male sterile tomatoes and afforded helpful information for understanding how RBOH genes regulating tomato reproduction process.