Improved native affinity purification of RNA.
RNA biochemical or structural studies often require an RNA sample that is chemically pure, and most protocols for its in vitro production use denaturing polyacrylamide gel electrophoresis to achieve this. Unfortunately, many RNAs do not quantitatively refold into an active conformation after denaturation, creating significant problems for downstream characterization or use. In addition, this traditional purification method is not amenable to studies demanding high-throughput RNA production. Recently, we presented the first general method for producing almost any RNA sequence that employs an affinity tag that is removed during the purification process. Because technical difficulties prevented application of this method to many RNAs, we have developed an improved version that utilizes a different activatable ribozyme and affinity tag that are considerably more robust, rapid, and broadly applicable.
Batey
Robert T
RT
Department of Chemistry and BioChemistry, University of Colorado, Boulder, CO 80309-0215, USA.
Kieft
Jeffrey S
JS
D000345:Affinity Labels / Q000737:chemistry; D001483:Base Sequence; D002846:Chromatography, Affinity / Q000379:methods*; D008969:Molecular Sequence Data; D009690:Nucleic Acid Conformation; D012313:RNA / Q000302:isolation & purification*; D016337:RNA, Catalytic / Q000302:isolation & purification
D000345:Affinity Labels; D016337:RNA, Catalytic; D012313:RNA