%0 Generic %A Abraham, Paul %A Adams, Rachel M. %A Tuskan, Gerald A. %A Hettich, Robert L. %D 2016 %T Moving Away from the Reference Genome: Evaluating a Peptide Sequencing Tagging Approach for Single Amino Acid Polymorphism Identifications in the Genus Populus %U https://acs.figshare.com/articles/dataset/Moving_Away_from_the_Reference_Genome_Evaluating_a_Peptide_Sequencing_Tagging_Approach_for_Single_Amino_Acid_Polymorphism_Identifications_in_the_Genus_i_Populus_i_/2391886 %R 10.1021/pr400192r.s013 %2 https://acs.figshare.com/ndownloader/files/4031575 %K 200 base pairs %K peptide sequence %K Peptide Sequencing Tagging Approach %K reference %K profile protein expression %K methionine %K approach %K oxidation %K Populu %K MS %K PST %K shotgun proteomics data %K energy collisional dissociation %K variant %K polymorphism %X The genetic diversity across natural populations of the model organism, Populus, is extensive, containing a single nucleotide polymorphism roughly every 200 base pairs. When deviations from the reference genome occur in coding regions, they can impact protein sequences. Rather than relying on a static reference database to profile protein expression, we employed a peptide sequence tagging (PST) approach capable of decoding the plasticity of the Populus proteome. Using shotgun proteomics data from two genotypes of P. trichocarpa, a tag-based approach enabled the detection of 6653 unexpected sequence variants. Through manual validation, our study investigated how the most abundant chemical modification (methionine oxidation) could masquerade as a sequence variant (Ala→Ser) when few site-determining ions existed. In fact, precise localization of an oxidation site for peptides with more than one potential placement was indeterminate for 70% of the MS/MS spectra. We demonstrate that additional fragment ions made available by high energy collisional dissociation enhances the robustness of the peptide sequence tagging approach (81% of oxidation events could be exclusively localized to a methionine). We are confident that augmenting fragmentation processes for a PST approach will further improve the identification of single amino acid polymorphism in Populus and potentially other species as well. %I ACS Publications