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Nucleic Acids Research, 1995, Vol. 23, No. 9 1570-1575
© 1995

METHODS

Increased stability of nucleic acids containing 7-deaza-guanosine and 7-deaza-adenosine may enable rapid DNA sequencing by matrix-assisted laser desorption mass spectrometry

Klaus Schneider and Brian T. Chait*

The Rockefeller University 1230 York Avenue, New York, NY 10021, USA

* To whom correspondence should be addressed

Received December 15, 1994. Revised March 16, 1995. Accepted March 16, 1995.

The use of matrix-assisted laser desorption mass spectrometry (MALDI-MS) has been suggested as an ultrafast readout of Sanger DNA sequencing ladders in a manner analogous to that used with sequencing gels. Currently, a serious limitation of MALDI-MS for the analysis of DNA results from the tendency for oligonucleotides to undergo facile fragmentation in the gas phase. The present study was undertaken to gain an understanding of the influence of various chemical structural features of purlne bases on the stability of oligodeoxynucleotide ions produced by MALDI. The study focused on the stability of model compounds of the type d(TTTTTTTTTTXTTTTTTTTTTTTT), where T designates deoxythymldlne and X a purine-contalning 2'-deoxynucleotide. A variety of different purine derivatives were chosen as the base in the nucleotlde X. The mass spectra of the model compounds containing 7-deaza analogues of guanine and adenlne reveal a significantly increased stability compared to the 7-aza analogues under the conditions of MALDI-MS. The previously reported incorporation of the 7-deaza-2'-deoxy-adenosine trlphosphate and the 7-deaza-2'-deoxy-guanosine triphosphate into DNA by polymerases suggests their use in a Sanger dideoxy sequencing experiment. The dideoxy termination products with the 7-deaza-purines instead of the 7-aza-purines might be sufficiently stable to allow separation and detection of the sequencing ladder by MALDI-MS. Thus, an ultrafast (seconds) read-out of DNA sequence may become feasible.


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