MS/MS spectra were searched against an in-house polypeptide sequence database corresponding to an improved annotation of the B. cereus ATCC 14,579 genome (Madeira et al., 2016a (link)). The MASCOT Daemon search engine (version 2.3.02; Matrix Science) was used to search tryptic peptides as previously described (Dupierris et al., 2009 (link); Madeira et al., 2016a (link)). The mass spectrometry proteomics data have been deposited in the ProteomeXchange Consortium (http://proteomecentral.proteomexchange.org ) via the PRIDE partner repository (http://www.ebi.ac.uk/pride ) with the dataset identifiers, PXD006169 and 10.6019/PXD006169 (exoproteome) and, PXD006205 and 10.6019/PXD006205 (cellular proteome).
Mascot daemon search engine
Manufactured by Matrix Science
Sourced in United Kingdom
The MASCOT Daemon search engine is a core component of the MASCOT suite of software tools developed by Matrix Science. It is designed to perform rapid and efficient protein identification from mass spectrometry data. The MASCOT Daemon is responsible for managing the search process and delivering results to users.
Automatically generated - may contain errors
4 protocols using mascot daemon search engine
1
Proteomic Analysis of B. cereus
2
Proteomic Analysis of BIS Protein
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
The BIS protein was immunoprecipitated from A172 cell lysates and separated by SDS–PAGE. Gels were stained with Instant Blue (Coomassie-based gel staining dye that is compatible with mass spectrometry analysis. Expedeon), and the visualized BIS band was excised. In-gel digestion was performed with modified porcine trypsin (Promega), and the peptides were analyzed by liquid chromatography-tandem mass spectrometry. The resulting peaks were searched against the SwissProt database (2014.07 release, 20210 sequences for human) using a Mascot Daemon search engine (version 2.4.0, Matrix Science), and the peptides of interest were quantified by peak area integration using the extracted ion chromatograms. We also predicted the phosphorylation motifs of BIS using the Motif Scan program24 (link) and Group-based Prediction System 2.0.25 (link)
3
Peptide Identification in C. jejuni
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
The peptides were enriched and separated using a lab-on-a-chip technology (Agilent, Massy, France) and fragmented using an on-line XCT mass spectrometer (Agilent). The fragmentation data were interpreted using the Data Analysis program (version 3.4, Bruker Daltonic, Billerica, MA, United States). For the protein identification, the MS/MS peak lists were extracted, converted into mgf-format files and compared to the C. jejuni, strain 81–176 protein database (UniprotKB, CP000538 for the chromosome, CP000549 for plasmid pTet and CP000550 for plasmid pVir) with the MASCOT Daemon search engine (version 2.6.0; Matrix Science, London, United Kingdom). The following search parameters were used: trypsin was used as the cutting enzyme, the mass tolerance for monoisotopic peptide window was set to ±1.0 Da and the MS/MS tolerance window was set to ±0.5 Da. Two missed cleavages were allowed. Carbamidomethylation, oxidized methionine, acetylation and pyroglutamate in Nt and amidation in Ct were chosen as variable modifications. Generally, the peptides with individual ions scores higher than the score indicated for p < 0.05 were selected. The proteins with two or more unique peptides matching the protein sequence were automatically considered as a positive identification. The main raw data are presented in Supplementary Data Sheet S1 . Other raw data are available upon request.
4
Comprehensive Proteogenomic Analysis of Bacillus cereus
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
An in-house polypeptide sequence database was made of the sequences of all previously annotated proteins encoded by the B. cereus ATCC 14579 chromosome (NC_004722) and the plasmid pBClin15 (NC_004721), and 44 proteins identified by a previous proteogenomic study (see Supplementary Table 1 in Ref. [28] ). This database, used to assign peptide sequences to MS/MS spectra, comprises 5299 polypeptide sequences totaling 1,464,675 amino acids. The MASCOT Daemon search engine (version 2.3.02; Matrix Science) was used for searching tryptic peptides with the following parameters: full-trypsin specificity, a mass tolerance of 5 ppm on the parent ion and 0.5 Da on the MS/MS, carboxyamidomethylated Cys (+57.0215) as a fixed modification and oxidized methionine (+15.9949) as a variable modification. The number of tolerated missed cleavages was set at 2. All peptide matches with a score below a p-value of 0.05 were filtered by the IRMa 1.28.0 parser [29] . A protein was considered validated when at least two different peptides were detected when considering all the samples. The falsepositive rate for protein identification was estimated using the appropriate decoy database as below 0.1% with these parameters. The mass spectrometry proteomics data have been deposited in the ProteomeXchange Consortium (http://proteomecentral.proteomeexchange.org) via the
About PubCompare
Our mission is to provide scientists with the largest repository of trustworthy protocols and intelligent analytical tools, thereby offering them extensive information to design robust protocols aimed at minimizing the risk of failures.
We believe that the most crucial aspect is to grant scientists access to a wide range of reliable sources and new useful tools that surpass human capabilities.
However, we trust in allowing scientists to determine how to construct their own protocols based on this information, as they are the experts in their field.
Ready to get started?
Sign up for free.
Registration takes 20 seconds.
Available from any computer
No download required
Revolutionizing how scientists
search and build protocols!