This article is published in Journal of Molecular Recognition as a special issue on Affinity 2009 edited by Gideon Fleminger, Tel-Aviv University, Tel-Aviv, Israel and George Ehrlich, Hoffmann-La Roche, Nutley, NJ, USA.
A new affinity approach to isolate Escherichia coli 6S RNA with histidine-chromatography†
Article first published online: 28 OCT 2010
Copyright © 2010 John Wiley & Sons, Ltd.
Journal of Molecular Recognition
Special Issue: Affinity 2009 – The 18th biennial meeting of the International Society for Molecular Recognition
Volume 23, Issue 6, pages 519–524, November/December 2010
How to Cite
Martins, R., Queiroz, J. A. and Sousa, F. (2010), A new affinity approach to isolate Escherichia coli 6S RNA with histidine-chromatography. J. Mol. Recognit., 23: 519–524. doi: 10.1002/jmr.1078
- Issue published online: 28 OCT 2010
- Article first published online: 28 OCT 2010
- Manuscript Accepted: 20 JUL 2010
- Manuscript Revised: 19 JUL 2010
- Manuscript Received: 13 NOV 2009
- affinity chromatography;
- RNA extraction;
- small RNA;
- 6S RNA
6S RNA is an abundant non-coding RNA in Escherichia coli (E. coli), but its function has not been discovered until recently. The first advance on 6S RNA function was the demonstration of its ability to bind the σ70-holoenzyme form of RNA polymerase, inhibiting its activity and consequently the transcription process. The growing interest in the investigation of non-coding small RNAs (sRNA) calls for the development of new methods for isolation and purification of RNA. This work presents an optimized RNA extraction procedure and describes a new affinity chromatography method using a histidine support to specifically purify 6S RNA from other E. coli sRNA species. The RNA extraction procedure was optimized, and a high yield was obtained in the separation of sRNA and ribosomal RNA (rRNA) from total RNA (RNAt). This improved method takes advantage of its simplicity and significant cost reduction, since some complex operations have been eliminated. A purification strategy was also developed to separate 6S RNA from an sRNA mixture. Pure RNA can be advantageously obtained using the histidine-affinity chromatography method, aiming at its application to structural or functional studies. Copyright © 2010 John Wiley & Sons, Ltd.