Chan et al. BMC Genomics 2011, 12:290 http://www.biomedcentral.com/1471-2164/12/290 We added both Group A and B proteins into Pepti- deAtlas, but only sections of the ORF as opposed to its entire length. Apart from the matched peptides, we had no further information to map the precise intron-exon boundaries. As a result, we only included the sequence spanned by the two outer-most peptide matches. In gene annotation, the most common form of mis- annotation is the incorrect placement of intron-exon boundaries, sometimes assigning a region as an intron where it should be an exon or vice versa. One example of a corrected (Group A) protein is in Figure 7a, where the peptide VQTVATPSIIER (PAp01421939, seven spec- tral counts) matched a contig (110756317+2_184592), from which protein XP_392304.3 was previously anno- tated. VQTVATPSIIER falls in the intronic region but the very high quality MS/MS spectra indicates that it is indeed real and that the intron-exon boundaries are mis-assigned for this protein. Gene annotation relies heavily on automated algo- rithms and pattern matching, which can sometimes com- pletely miss real genes. Figure 7b shows an example a novel (Group B) protein - an ORF with several high-qual- ity matches from new peptides. A BLAST search against the non-redundant database revealed no significant hits, which is not surprising, given that gene annotation algo- rithms often rely on sequence similarity against other organisms and so if there had been a hit in another organism, this gene might have been identified as such. It should be noted that since most bee genome contigs have not be scaffolded, it is possible that one protein may span multiple contigs; thus, some of the novel, expressed ORFs detected here may come from the same protein. In order to facilitate analysis of future honey bee shotgun experiments, we have compiled a spectral library based on all the identifications in the PeptideA- tlas build using the SpectraST library building tool [31]. SpectraST collects all replicate spectra for each peptide ion and creates a consensus spectrum based on a voting scheme that retains repeated peaks. This can enhance future experiments because SpectraST searches are many times faster than conventional sequence searching and SpectraST scores can discrimi- nate better between correct and incorrect identifica- tions [32]. The spectrum library in splib and sptxt format is available at the same URL with the other data products from this work. Targeted proteomics workflows via selected reaction monitoring (SRM) enable highly sensitive and repeatable quantitative measurements on triple-quadrupole mass spectrometers [33]. However, such workflows require considerable experiment planning to select the specific signatures needed to detect the target peptides [34]. To Page 9 of 11 aid in this experiment planning, we have created an SRMAtlas build [35] based on the ion trap observations and predictions. The results of this process are available at the SRMAtlas web site http://www.srmatlas.org. Conclusions We present here the first publicly accessible resource for honey bee proteomics using the PeptideAtlas architec- ture [7]. In addition to providing the experimental evi- dence behind each peptide identification, we have also undertaken a proteogenomic re-annotation of honey bee proteins that has led to the identification of 186 new or mis-annotated regions of bee proteins. We expect that as more MS/MS data are collected, we will be able to further refine the annotation of honey bee genes. Honey bees are typically studied to reveal the biologi- cal underpinnings of a complex insect society and rarely as a model of human disease or basic biology. Conse- quently, there is a dearth of detection reagents and probes for honey bee proteins. As we and others move into using selected reaction monitoring as a means for targeting specific proteins, the MS/MS data presented in the Honey Bee PeptideAtlas will provide the empirical evidence required to make intelligent decisions about the design of these experiments. The importance of honey bee populations to natural and agro-ecological systems, coupled to their current decline obligates us to improve the status quo. The release of these data into the public domain not only represents validation and improvement of the current annotation of the reference genome, but provides the empirical evidence to guide future honey bee biochemical research. Availability and Requirements Project name: HoneyBee PeptideAtlas Project home page: https://db.systemsbiology.net/ sbeams/cgi/PeptideAtlas/buildDetails? atlas_build_id=282 Operating system(s): e.g. Platform independent Programming language: Other requirements: none License: Creative Commons Attribution Any restrictions to use by non-academics: none Additional material Additional file 1: Correct or novel honey bee proteins supported by MS-based peptide spectra. Peptide spectra were searched against six frame-translated honey bee ORFs to augment currently existing, publically available protein databases. Each protein in this file is hyperlinked to the relevant entry in PeptideAtlas, which provides a graphical display of MS-based peptide evidence for each Corrected (Column A) or Novel Protein (Column B).