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Polystyrene reverse-phase ion-pair chromatography of chimeric ribozymes
P M Swiderski1, E L Bertrand, B E Kaplan
1Beckman Research Institute of the City of Hope, Duarte, California 91010.
Analytical Biochemistry
|January 1, 1994
Summary
A new reverse-phase ion-pair HPLC method using polystyrene resin efficiently purifies synthetic DNA-RNA oligomers. This faster, reliable technique leverages ion pair formation for rapid separation of chimeric ribozymes.
Area of Science:
- Biochemistry
- Molecular Biology
- Organic Chemistry
Background:
- Synthetic chimeric DNA-RNA oligomers are crucial for research.
- Existing purification methods for these molecules can be slow and unreliable.
- Efficient purification is essential for downstream applications.
Purpose of the Study:
- To develop a faster and more reliable method for purifying large quantities of synthetic chimeric DNA-RNA oligomers.
- To utilize reverse-phase ion-pair High-Performance Liquid Chromatography (HPLC) on a polystyrene resin for this purpose.
Main Methods:
- Employed a reverse-phase polystyrene resin for ion-pair HPLC.
- Utilized tetrabutylammonium fluoride in the final cleavage step of synthetic oligoribonucleotides.
- Exploited the serendipitous formation of ion pairs between tetrabutylammonium cations and oligomer phosphates.
Main Results:
- The ion pair formation retarded oligomer elution during HPLC.
- This retardation enabled rapid removal of excess tetrabutylammonium fluoride.
- Achieved concomitant purification of synthesized chimeric ribozymes.
- Demonstrated a correlation between ion-paired oligomer length and HPLC retardation.
Conclusions:
- Developed a novel, efficient, and reliable reverse-phase ion-pair HPLC method for purifying synthetic chimeric DNA-RNA oligomers.
- The technique offers significant advantages in speed and reliability over previous methods.
- Successfully applied the method to the purification of synthesized chimeric ribozymes.