VIGS of CaNAC2c/d and CaHSFA5 in pepper plants
To silence CaNAC2c(d) or CaHSFA5 in pepper plants by virus induced gene silencing (VIGS), GV3101 cells containing PYL192 and PYL279-CaNAC2c (CaNAC2d , CaHSFA5 ), or PYL279 (as a negative control) were mixed and resuspended in the induction medium at 1:1 ratio to OD600 = 0.6, which were infiltrated into cotyledons of 2-week-old pepper plants. To simultaneously silenceCaNAC2c and CaNAC2d in pepper plants, GV3101 cells containing PYL192 and PYL279-CaNAC2c and cells containing PYL192 and PYL279-CaNAC2d were mixed at ratio 1:1 and resuspended in the induction medium to OD600 = 0.6. 100 μl were infiltrated into cotyledons of 2-week-old pepper plants, which were placed in dark at 16 ℃ for 56 h, and then moved to a growth room under conditions of 28 ℃, 60–70 mmol photons m-2 s-1, a relative humidity of 70%, and a 16-h light/8-h dark. At 15 days post inoculation (dpi), the success and specificity of gene silencing were assayed in pepper plants challenged with HTS by measuring the transcript level of CaNAC2c (CaNAC2d , CaHSFA5 ) or the transcript level of CaNAC2c and CaNAC2d .