Gut, I. G., Jeffery, W. A., Pappin, D. J. C., Beck, S. (1997) Analysis of DNA by 'charge tagging' and matrix-assisted laser desorption/ionization mass spectrometry. Rapid Communications in Mass Spectrometry, 11 (1). pp. 43-50. ISSN 09514198 (ISSN)
Abstract
We have developed a method to quantitatively attach quarternary ammonium fixed charge tags to the 5' or 3'NH2 ends of DNA using N-hydroxysuccinimidyl ester chemistry. The chemical conditions for tagging were chosen so that tagging takes place exclusively on aliphatic NH2 groups while base amino groups remain unmodified. The charge tagging chemistry was combined with a previously developed backbone alkylation procedure for phosphorothioate DNA. The efficiency of the detection in matrix-assisted laser desorption/ionization (MALDI) mass spectrometry of unmodified and modified DNA (phosphorothioate backbone, charge tagged, backbone alkylated, and charge tagged and backbone alkylated) was investigated using a series of different matrices. For a-cyano-4-hydroxycinnamic acid (a matrix, commonly used for the analysis of proteins, but which gives unsatisfactory results with unmodified DNA). For instance, the charge tagged and backbone alkylated DNA is detectable with a sensitivity and resolution comparable with that for peptides. The combination of charge tagging and backbone alkylation with the use of a suitable matrix improves the detectability of small oligonucleotides by MALDI by a factor greater than 100 compared to unmodified oligonucleotides.
| Item Type: | Paper |
|---|---|
| Subjects: | bioinformatics > genomics and proteomics > genetics & nucleic acid processing > DNA, RNA structure, function, modification Investigative techniques and equipment > spectroscopy > mass spectrometry |
| CSHL Authors: | |
| Communities: | CSHL labs > Pappin lab |
| Depositing User: | Kathleen Darby |
| Date: | 1997 |
| Date Deposited: | 08 May 2014 18:00 |
| Last Modified: | 24 Feb 2017 16:48 |
| URI: | https://repository.cshl.edu/id/eprint/29963 |
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