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A revisit of high collision energy effects on collision-induced dissociation spectra using matrix-assisted laser desorption/ionization tandem time-of-flight mass spectrometry (MALDI-LIFT-TOF/TOF): application to the sequencing of RNA/DNA chimeras


  • Florence Mauger,

    1. CEA/Institut de Génomique/Centre National de Génotypage, Evry Cedex, France
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  • Jean-Claude Tabet,

    1. UPMC/IPCM (Chimie Biologique Organique et Structurale), UMR 7201, Paris Cedex 05, France
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  • Ivo G. Gut

    Corresponding author
    1. CEA/Institut de Génomique/Centre National de Génotypage, Evry Cedex, France
    2. Centro Nacional de Análisis Genómico, Barcelona, Spain
    • Correspondence to: I. G. Gut, CEA/Institut de Génomique/Centre National de Génotypage, Bâtiment G2, 2 rue Gaston Crémieux, CP 5721, 91057 Evry Cedex, France.


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High-energy collision-induced dissociation (CID) spectra of isomeric RNA/DNA chimeras using matrix-assisted laser desorption/ionization time-of-flight LIFT mass spectrometry (MALDI-LIFT-TOF/TOF) can potentially be applied for an exhaustive fragment characterization in a nucleic acid sequencing scheme. These chimeras contain deoxynucleotides and at the 3'-end a ribonucleotide with a 3'-phosphate group.


Deprotonated RNA/DNA chimeras of 4-, 5-, 7- and 10-mers are analyzed by CID. This enhances consecutive dissociations from both the precursor and prompt product anions generated by MALDI and metastable fragmentations prior to entering the LIFT cell.


Gas-phase fragmentations of 4- and 5-mers produced many fragment ions, from base release prior to consecutive cleavage of the nucleotide phosphate bond linkage phosphate. The unusual a4 product ion is a specific and diagnostic dissociation of the 4-mer if the ribonucleotide contains cytosine. As the size of RNA/DNA chimeras increase, several abundant product ions are generated mainly from zwitterionic forms (deprotonated phosphate ester and protonated base sites): [(M-H)–BiH], [ai–BiH], wj, [wj, (ai-BiH)] (if Bi ≠ T) as internal product ion, and more rarely [wj–BiH]. The absence of the majority of the [ai–BiH] series although the wj series suggested that the higher critical energy processes with a loose transition state are favored yielding the wj series. A large number of abundant fragment ions are detected which enable each isomer to be sequenced.


This sequencing method is high-throughput, accurate and could be used to sequence isomers of up to 10-mers and also oligonucleotides of unknown sequence. However, RNA/DNA chimeras without thymine must be sufficiently concentrated to reach desorption of deprotonated molecular species to be selected in LIFT to produce all fragment ions within measurable abundances. Copyright © 2014 John Wiley & Sons, Ltd.