Rapid Creation of New Photoperiod-/Thermo-Sensitive Genic Male-Sterile Rice Materials by CRISPR/Cas9 System

SUMMARY Rice is an important food crop in China, and the development of hybrid rice is a crucial way to increase grain yield. The creation of dual-purpose nuclear-sterile lines for two-line hybrid breeding has become vital for commercial rice breeding. We constructed the pC1300-2x35S::Cas9-sgRNA PTGMS2-1 expression vector for editing the male fertility gene PTGMS2-1 in two widely compatible rice varieties, 93-11 and Huazhan, by using the CRISPR/Cas9 system. We obtained the marker-free photoperiod-/thermo- sensitive genic male-sterile (P/TGMS) lines in T 1 generation. According to the experiments in phytotron with four temperature and photoperiod treatments, we found the temperature is the main factor for restoring the pollen fertility of ptgms2-1 mutants in 93-11 and Huazhan, and the photoperiod also has some effects on pollen fertility in two different rice backgrounds. The application of cultivating new male-sterile lines by genome editing system will significantly accelerate the rice breeding process.