In vitro transcription of c-jun gene using fractionated nuclear extract from regenerating rat liver

Regenerating rat liver serves as a source of proliferating cells, such a system can be used to study the regulation of genes involved in proliferation. We have established an in vitro transcription capable of supporting accurate transcription of cloned c-jun gene using fractionated nuclear extract prepared from partially hepatectomized rat liver. EcoR I linearized plasmid -132/+170 jun-CAT containing c-jun gene promoter region (-132 to +170 nucleotides) was transcribed in an in vitro run-off transcription assay and a transcript of expected size i.e. 370 nucleotides was obtained. The intensity of the transcript was dependent on the concentration of fractionated nuclear extract as well as template. The transcription was alpha-amanitin sensitive indicating that it is directed by RNA polymerase II.