Reassessing the contribution of TOMATO AGAMOUS-LIKE1 to fruit ripening by CRISPR/Cas9 mutagenesis

Key message CRISPR/Cas9-edited TOMATO AGAMOUS-LIKE1 ( TAGL1 ) provided new insights into fruit ripening . Abstract TOMATO AGAMOUS LIKE 1 (TAGL1) has been identified as playing a key role in the process of tomato fruit development and ripening. We have re-evaluated the functions of TAGL1 using CRISPR/Cas9 mutagenesis. Three KO mutants contained frameshift mutations resulting in premature termination codons due to a 1 bp insertion. TAGL1 -KO mutants exhibited dark immature fruits and orange ripening fruits. The fruit shape was characterized by a prominent pointed tip at the end and the pericarp thickness was significantly thinner. TAGL1 -KO mutants showed reduced ethylene biosynthesis, increased firmness, and delayed onset of ripening. The chlorophyll content of TAGL1 -KO mutants was higher in the mature green stage and the lycopene content of TAGL1 -KO mutants in the ripening stage was lower compared to the WT. ACS2 , ACS4 , ACO1 , ACO3 , PG2a , PL , PME , EXP1 , and PSY1 in the mutants were significantly down-regulated during ripening. Ripening fruits in the double mutant of rin and tagl1 showed a more extreme phenotype than the rin mutant suggesting that the double mutation acts synergistically during ripening. TAGL1-targeted mutagenesis by CRISPR/Cas9 strengthens its regulatory functions controlling ripening parameters and provides new insights into fruit ripening.