In vitro coupled transcription splicing.
Many convincing studies published in recent years strongly support coupling of transcription and pre-mRNA processing. Despite key advances in our understanding of these processes, there is a lack of a robust in vitro system in which to study the mechanism of this coupling for complex pre-mRNAs. Here, we describe an in vitro system capable of transcribing and splicing complex transcripts with three and four exons. We also demonstrate how the system can be used to study exon silencing in vitro. We believe that this system will be a useful tool to study the mechanisms that mediate the coupling of transcription and pre-mRNA processing.
Natalizio
Barbara J
BJ
Department of Molecular Genetics, Center for RNA Biology, Duke University Medical Center, Durham, NC 27710, USA.
Garcia-Blanco
Mariano A
MA
D012321:DNA-Directed RNA Polymerases / Q000235:genetics; D003851:Deoxyribonucleases; D005091:Exons; D005914:Globins / Q000096:biosynthesis; D006801:Humans; D008967:Molecular Biology / Q000379:methods*; D034622:RNA Interference; D012319:RNA Polymerase II / Q000235:genetics; D012322:RNA Precursors / Q000502:physiology*; D012326:RNA Splicing; D013696:Temperature; D014158:Transcription, Genetic; D014764:Viral Proteins / Q000235:genetics
D012322:RNA Precursors; D014764:Viral Proteins; D005914:Globins; D012319:RNA Polymerase II; C062210:bacteriophage T7 RNA polymerase; D012321:DNA-Directed RNA Polymerases; D003851:Deoxyribonucleases