Fig. 4

BMV VIGS is more efficient when an anti-sense strand of the gene fragment is used for silencing. Two-week old sorghum plants were rub inoculated with BMV containing sap of N. benthamiana. a The BMV::Ubiq and BMV::anti-Ubiq infected plants had a varying degree of Ubiq silencing. The extent of silencing was more severe in the BMV::anti-Ubiq infected plants. The experiment was repeated three times with similar results. b BMV level was analyzed by western blot using an antibody against BMV coat protein. BMV level was more in the sorghum plants infected with BMV: anti-Ubiq compared to BMV::Ubiq. c RT-PCR was performed to detect the presence of gene fragment in the BMV RNA3 using primers flanking the insert. The BMV with insert was present in the sorghum infected with BMV::anti-Ubiq but not detectable in sorghum infected with BMV::Ubiq. For RT-PCR and western blot, leaves of three N. benthamiana plants or 20 sorghum plants per sample were pooled to extract the virus