TEST OF THE POTENTIAL OF A DATP SURROGATE FOR SEQUENCING VIA MALDI-MS

1-(2'-Deoxy-beta-D-ribofuranosyl)-3-nitropyrrole phosphate was incorpo rated into a DNA decamer and analyzed via matrix-assisted laser desorp tion ionization mass spectrometry (MALDI-MS). The extent and compositi on of the various fragment peaks were compared with those in the MALDI -MS spectrum of dT(4)AT(5), The nitropyrrole-containing oligomer prove d to be more robust, Two different DNA template assays were then used to attempt to identify DNA replicating enzymes that would incorporate the corresponding triphosphate, i.e. 1-(2'-deoxy-beta-D-ribofuranosyl) -3-nitropyrrole triphosphate (dXTP), It was shown that dXTP was not in corporated by some enzymes and it inhibited others, However, DNA polym erase I Klenow fragment and avian myeloblastosis virus reverse transcr iptase incorporated dXTP in place of dATP and then replicated the temp late overhang in the usual way, The potential of dXTP as a surrogate f or dATP in DNA sequencing with MALDI-MS analysis is discussed.