each with 5 of the electroporated cells at 3 48 and 72 h post electroporation the cells were harvested and assayed for luciferease activity promega madison wis the luciferase levels at 3 h post electroporation were used to correct for transfection efficiency huh 7 cells were transfected with pcep4d plasmids expressing sirna 6367 sirna 6367 mm or with no insert the plasmids were transfected into huh 7 cells using lipofectamine2000 invitrogen carlsbad calif and suggested method medium containing 75 u03bcg ml hygromycin invitrogen carlsbad calif was added to the cells 24 h post transfection rna samples were purified from huh 7 cells using trizol reagent life technologies invitrogen carlsbad calif total rna 5 u03bcg was treated with glyoxal and subjected to electrophoresis in a 0 9 agarose gel using standard techniques 31 the gels were transferred to hybond n nylon membrane amersham pharmacia piscataway n j and probed with 32 p labeled neomycin resistance gene dna which had been labeled using the ready to go dna labeling kit amersham pharmacia piscataway n j an hcv sense strand specific riboprobe was made using hind iii linearized phcvrepab12 replicon plasmid as a template for use in the riboprobe t7 system with u03b1 32 p utp promega madison wis equal numbers of naive or replicon containing huh 7 cells were lysed in sds sample buffer 72 h after electroporation with sirnas protein was electophoresed on.