A single-base deletion in ZjSGR confers the STAY-GREEN phenotype and the enhanced stress tolerance in Zoysia japonica under dark and low temperature stresses

Zoysia japonica (Zoysia japonica Steud.) is a perennial warm-season grass widely used for soil and water conservation due to its extensive root system and resilience in lands, which has a huge potential in enhancing carbon sinks, restoring degraded ecosystems, and improving the living environment. However, Z. japonica will lose color and go into withering prematurely in late fall and winter compared with cool-season grass in the transition zone and the northern regions, which limits the application of Z. japonica. In this study, we successfully cultivated a “G” single-base deletion mutant of ZjSGR (STAY-GREEN) in Z. japonica (sgr-mutant) by CRISPR/Cas9 transient expression without exogenous genes. Based on the experiments of the sgr-mutant and the wild-type plant (WT-plant) under dark and low temperature stresses, the study demonstrates that CRISPR/Cas9-mediated novel single-base deletion of ZjSGR in Z. japonica exhibits not only prolonged chlorophyll retention but also superior stress tolerance under both dark and low temperature stress treatments. The mechanism of the improved stress tolerance of the sgr-mutant includes the accumulation of chlorophyll and carotenoid, providing a foundational buffer against oxidative damage, and a flexible regulation of key antioxidant enzymes in an energy-efficient way with a lower transcriptional burden compared to the WT-plant. Therefore, our study demonstrates that SGR plays a vital role in both leaf color and stress tolerance in grass, providing a new breeding strategy for developing green-persistence, stress-tolerance and low-maintenance grass applications.