Modulation of fruit softening by antisense suppression of endo-beta-1,4-glucanase in strawberry

Abstract

To test the effect of endo-beta-1,4-glucanase (EGase) in strawberry, we produced transgenic strawberry plants that contained sense and antisense cDNA encoding strawberry EGase under the CaMV 35S promoter (CaMV35S-P) and the strawberry fruit dominant ascorbate peroxidase (APX) promoter. Independent transgenic lines were generated and the firmness of the fruit was characterized after harvest. Interestingly, transgenic lines that harbored the cDNA antisense under the CaMV 35S promoter were not generated, but transgenic lines that contained sense EGase cDNA (FraCel1) under the CaMV 35S promoter and sense and antisense EGase cDNA under the APX promoter were successfully obtained using a tissue culture system. Reverse transcriptase polymerase chain reaction (RT-PCR) analysis revealed that the steady-state transcript levels of CaMV35S-P:FraCel1 sense transgenic lines were dramatically increased in fruit evaluated at all stages, as well as in the leaves. Moreover, real-time PCR analysis demonstrated that the steady-state transcript level of FraCel1 in independent transgenic lines that contained antisense FraCel1 under the APX promoter was reduced in fruit during the turning and red stages when compared with wild-type strawberries. These results were consistent with the firmness of strawberries that contained the antisense FraCel1 under the APX promoter, which was 19-36% greater in the turning stage and 22-25% greater in the red stage when compared to wild-type strawberries. Taken together, these findings indicate that fruit-specific down-regulation of the EGase gene using the APX promoter is an effective technique for increasing fruit firmness in strawberries.