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JoVE Journal Biochemistry
Single-step Purification of Macromolecular Complexes Using RNA Attached to Biotin and a Photo-cleavable Linker
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Single-step Purification of Macromolecular Complexes Using RNA Attached to Biotin and a Photo-cleavable Linker

Article DOI: 10.3791/58697-v • 08:12 min • January 3rd, 2019
January 3rd, 2019 •
1, 1, 1,2
1Integrative Program for Biological and Genome Sciences, University of North Carolina at Chapel Hill, 2Department of Biochemistry and Biophysics, University of North Carolina at Chapel Hill

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Summary
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RNA/protein complexes purified using botin-streptavidin strategy are eluted to solution under denaturing conditions in a form unsuitable for further purification and functional analysis. Here, we describe a modification of this strategy that utilizes a photo-cleavable linker in RNA and a gentle UV-elution step, yielding native and fully functional RNA/protein complexes.

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RNA Protein Complexes Purification Mass Spectrometry Single-step Purification Limited Amounts Dissociation Multi-step Purification Procedures RNA Binding Sites Xiao-cui Yang T7 Generated RNA Adaptor PCB Oligonucleotide Binding Buffer Boiling Water Room Temperature Mouse Nuclear Extract EDTA Tagged RNA Substrate PCB Moiety Incubate On Ice Microcentrifuge
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