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Nucleic acid mass spectrometry - UAB

2/2/20111 Nucleic acid and nucleotide mass spectrometryStephen Barnes, 744 02-02-11 classSynopsis MALDI Sample prep UV laser SNPs, multiplexing IR laser (large DNA/RNA) ESI Bases Nucleosides Nucleotides2/2/20112UV-MALDI TOF of Nucleic acids Limited in the number of residues (up to 70 nt)12 nt Antisense Oligo DNAF erulic acid as the matrix254 nm laserKeough et al., 1996 Single nucleotide polymorphisms (SNPs)2/2/20113 Multiplexing concept for SNPsATCTGAAACGGTCACAGACCTGGAB iotinTTTTTTTTTGATCTGAAACGGTCACAACCTGGCB iotinTTTTTTTTTTTTTCTGAAACGGTCACAACCTGGGA GB iotinTTTTTTTTTTTTTTD .013 Cytosine MALDI-TOF spectra Mengel-J rgensen et al., Anal Chem 200417-plex analysis2/2/20114 Infra-red MALDI-TOF of DNAB erkenkamp et al., Science 1998300 fmol, 1 shot300 amol, 30 shots516 nt PCR productPptd with EtOH from 2 M ammonium acetateIR laser at mmSample in glycerolIR MALDI of Nucleic acidsPptd with EtOH from 2 M ammonium acetateIR laser at mmSample in glycerololigoBerkenkamp et al.

2/2/2011 5 Fundamentals of nucleic acids analysis by ESI-mass spectrometry •Analysis as bases, nucleosides –No different from other small molecules –Bases are sufficiently hydrophobic to be analyzed by reverse-phase LC-MS •The addition of ribose or deoxyribose not a problem •Detected in the positive ion mode •Nucleotides are more …

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