In vitro synthesis of dsRNA complementary to GFP (dsGFP)

PCR products for in vitro synthesis of dsGFP were generated from plasmid templates using primers containing the T7 polymerase recognition sequence in their 5’-ends. PCR products were purified using Phenol:Chloroform:Isoamyl Alcohol (25:24:1 v/v). dsRNAs were synthesized from purified PCR products using MegaScript T7 transcription kit (Invitrogen), purified using Acid Phenol (Invitrogen) and annealed (20mM Tris-HCl pH7.5, 100mM NaCl). Size and annealing efficiency were evaluated in 2% agarose gel stained with Ethidium Bromide.