Systematic mutagenesis reveals dominant–minor paralog configurations in the rice GA2ox gene family

Introduction Gibberellin 2-oxidases (GA2oxs) deactivate bioactive gibberellins and play essential roles in regulating rice growth and development. However, the relative contributions of individual OsGA2ox paralogs, their redundancy patterns, and dominant–minor relationships within this multigene family remain poorly understood. Clarifying these relationships is important for understanding how GA2ox genes coordinately influences plant development in rice. Methods To address these questions, we generated 36 CRISPR/Cas9-edited rice lines targeting all nine OsGA2ox genes, including single-, double-, triple-, and quadruple-gene knockout mutants, and systematically analyzed their effects on agronomic traits to compare gene-specific and combinatorial functions. Results In Class II, OsGA2ox1 is the main regulator of plant height, whereas OsGA2ox2 alone showed no discernable effect. Double mutants of these two genes showed a modest height increase and a reduced grain number but retained normal flowering time, indicating partial redundancy in regulating plant height and grain number, with OsGA2ox1 as the dominant contributor. In Class I, OsGA2ox3 , OsGA2ox4 , and OsGA2ox8 function as dominant suppressors of stem elongation, whereas OsGA2ox7 showed no phenotypical effect in single mutants but contributed to stem elongation control when disrupted together with other Class I members. The quadruple Class I mutant exhibited the greatest increase in plant height but reduced grain weight and fertility. In Class III, OsGA2ox5 and OsGA2ox6 regulated plant height, with OsGA2ox6 additionally influencing grain size, whereas OsGA2ox9 showed no detectable function. Discussion These findings reveal a hierarchical but partially redundant regulatory framework within the rice GA2ox gene family, in which dominant and minor paralogs contribute unequally to GA-related growth regulation. Our study provides allele-validated genetic evidence for functional diversification among OsGA2ox paralogs and highlights how redundancy and specialization are integrated to stabilize hormone-regulated development and key agronomic traits in rice.