title: Biotin-Streptavidin Affinity Purification of RNA-Protein Complexes Assembled In Vitro.
abstract: RNA-protein complexes are essential for the function of different RNAs, yet purification of specific RNA-protein complexes can be complicated and is a major obstacle in understanding the mechanism of regulatory RNAs. Here we present a protocol to purify RNA-protein complexes assembled in vitro based on biotin-streptavidin affinity. In vitro transcribed RNA is labeled with (32)P and biotin, ribonucleoprotein particles or RNPs are assembled by incubation of RNA in nuclear extract and fractionated using gel filtration, and RNP fractions are pooled for biotin-streptavidin affinity purification. The amount of RNA-protein complexes purified following this protocol is sufficient for mass spectrometry.
authors.affiliation: Institute of Cancer Stem Cell, Cancer Center, Dalian Medical University, No. 9 West Section Lvshun South Road, Dalian, Liaoning, 116044, People's Republic of China.
authors.forename: Shuai
authors.initials: S
authors.lastname: Hou
authors.affiliation: Institute of Cancer Stem Cell, Cancer Center, Dalian Medical University, No. 9 West Section Lvshun South Road, Dalian, Liaoning, 116044, People's Republic of China.
authors.forename: Lei
authors.initials: L
authors.lastname: Shi
authors.affiliation: Institute of Cancer Stem Cell, Cancer Center, Dalian Medical University, No. 9 West Section Lvshun South Road, Dalian, Liaoning, 116044, People's Republic of China. haixinlei@dlmedu.edu.cn.
authors.forename: Haixin
authors.initials: H
authors.lastname: Lei
mesh_terms: D001710:Biotin / Q000737:chemistry*; D002846:Chromatography, Affinity / Q000379:methods*; D013058:Mass Spectrometry / Q000379:methods; D012313:RNA / Q000302:isolation & purification; D012261:Ribonucleoproteins / Q000302:isolation & purification*; D019809:Streptavidin / Q000737:chemistry*
chemical_list: D012261:Ribonucleoproteins; D012313:RNA; D001710:Biotin; D019809:Streptavidin
keywords: Affinity purification; Biotin; RNA–protein complexes; Streptavidin