In vitro transcription of a cloned vaccinia virus gene by a soluble extract prepared from vaccinia virus-infected HeLa cells

Abstract

Faithful transcription of a vaccinia virus gene was accomplished in vitro by using a soluble extract prepared from vaccinia virus-infected HeLa cells. Specific transcription of the cloned vaccinia virus gene was detected by using template DNA restricted within the transcribed region. The vaccinia virus gene was not transcribed by extracts prepared from uninfected HeLa cells even with supplementation by purified vaccinia virus RNA polymerase, nor was a clone of adenovirus 2 DNA bearing the major late promoter transcribed by the extract from vaccinia virus-infected HeLa cells. Thus, infection by vaccinia virus altered cellular transcriptional specificity to favor expression of vaccinia virus genes. RNA synthesis by the infected cell extract was resistant to alpha-amanitin but strongly inhibited by beta, gamma-imido ATP and novobiocin.