Directional antisense and sense cDNA cloning using Epstein-Barr virus episomal expression vectors

Abstract A set of Epstein-Barr virus (EBV) episomal expression vectors, incorporating either the Rous sarcoma virus 3' long terminal repeat or the human metallothionein II A gene promoter, were constructed. The transcriptional cassettes encompassed by these vectors were designed to permit both antisense and sense RNA transcription. A novel methodology was developed for directional cDNA cloning using an oligodeoxyribonucleotide adapter; the EBV episomal vectors alternatively enabled the insertion of cDNA segments in antisense or sense orientations. We propose a strategy for random antisense RNA mutagenesis exploiting this vector system and a method for episome-based directional antisense cDNA cloning and expression, permitting the rapid identification of genes mediating selectable cellular functions.