title: In vitro analysis of transcriptional activators and polyadenylation.
abstract: In vitro assays have provided a valuable tool to study the mechanism of 3' processing of eukaryotic mRNA precursors and have contributed a great deal to the identification of factors that carry out and regulate 3' processing. Previously, we have shown that transcriptional activators directly enhance polyadenylation by utilizing in vitro transcription-coupled polyadenylation with the prototypical transcription activator GAL4-VP16. In this chapter, we describe a detailed protocol for this assay, which will be useful in examining potential roles for other transcription-related factors in 3' processing and other questions related to the coupling of transcription and mRNA polyadenylation.
authors.affiliation: Biomedical Research Institute, National Institute of Advanced Industrial Science and Technology, Tsukuba-shi, Ibaraki, Japan.
authors.forename: Takashi
authors.initials: T
authors.lastname: Nagaike
authors.forename: James L
authors.initials: JL
authors.lastname: Manley
mesh_terms: D026723:Polyadenylation / Q000502:physiology*; D012333:RNA, Messenger / Q000378:metabolism*; D015534:Trans-Activators / Q000378:metabolism*
chemical_list: C070633:Gal-VP16; D012333:RNA, Messenger; D015534:Trans-Activators