Validation of the bHLH gene (
) with virus-induced gene silencing. A. Cotyledon agroinfiltration of TRV vectors was carried out in the S plantlets at the cotyledon stage. R plants treated with Phytoene desaturase (PDS) gene silencing constructs pTRV1 and pTRV2 - (PDS) showed bleached areas in leaflets (left). S plants treated with pTRV1 and pTRV2 vectors showed the normal phenotype (middle). S plantlets treated with SlybHLH131 gene silencing constructs pTRV1 and pTRV2 - (SlybHLH131) showed a cell death phenotype (right). Arrow indicates cell death. B. Trypan blue staining of VIGS and empty vector lines leaves. C. DAB staining of VIGS and empty vector lines leaves. D. Relative expression of SlybHLH131 transcript using real-time RT-PCR analysis in the VIGS-treated lines 20 days after agroinfiltration with TRV vectors. Tomato α-Tubulin (Solyc04g077020.2) was used as an internal reference. Error bars represented SE of three biological replicates and asterisks indicate significant differences by Student’s t test for P < 0.05.