A General Method for Isolating Targets of RNA and DNA Binding Proteins
- Comments & Ratings
To isolate and characterize in vivo targets of RNA and DNA binding proteins, our laboratory used a novel strategy in which biotinylated protein produced using TnT® coupled transcription/translation system is bound to a softlink™ soft release avidin solid support and used to capture either mrna (from total rna) or dna. bound nucleic acids are eluted from the solid support and then amplified by differential display rt-pcr (ddrt-pcr). bands specific for the target protein are reamplified and cloned into the pgem®-t vector for subsequent sequencing. cloned products can then be used to define the binding motif in the target using a sensitive biotinylated protein capture assay
Promega Notes 67, 5.
Robert B. Denman
Department of Molecular Biology, Laboratory of Molecular Neurobiology, New York State Institute for Basic Research in Developmental Disabilities, NY
Publication Date: 1998