Biotin-Enhanced Fragmentation for Direct Deoxyribo-Nucleic Acid Sequencing Using Matrix-Assisted Laser Desorption/Ionization Mass Spectrometry

Fragmentation of synthetic oligonucleotides under the influence of biotin was investigated using 3-hydroxypicolinic acid (3-HPA) as a matrix-assisted laser desorption/ionization (MALDI) matrix. Addition of biotin into the sample enhanced fragmentation of the oligonucleotide between bases. However, when the biotin was tagged to the 5′-terminus of the oligonucleotide, enhancements were observed not only in desorption/ionization efficiency but also in the fragmentation of molecular ions. The fact that the protonation/deprotonation process occurs on the tagged biotin is a possible reason for the enhancement in desorption/ionization. Site-specific backbone cleavage fragmentation patterns were observed. The sequences of oligonucleotides can be obtained from their fragment ions. The direct sequencing of a 5′-biotin-tagged 25-mer is demonstrated.