User Manual-Peptide Depot
Contents
1. Configure DSN according to your mysql server info Choose sequest as the database Youare almost all set Open peptide depot in FileMaker and go to menu file gt manage gt external data sources Select Peptide_Depot_SQL and type in username as password this is optional if password is not set FileMaker will ask you password every time you login to this database Now your FileMaker and MySQL database should be all linked up Try it out and tell us what you think Navigating Peptide Depot Peptide Depot Home Page This page is this first page you will see when you open up peptide depot It acts as a portal to other views in the database From here you can access all experiments and experiment comparisons in the database As analysis of experiments and comparisons is the main functionality of this software this is probably what the homepage will be used for most of the time However buttons at the bottom will lead you to other features of the software Peptide Depot Currently Logged in as nhung Log off of windows to login as different user Experiments Collection YiYuan Subcollection Jurkat 201 3rd Experiment jurkat 2D1 3rd 1 Sm 01 07130 New Experiment Load Comparisons mma news Comparison JurkatSip76041709_mascot Load Other Tools Q 75 maha Browse Kryan lesung Change Log Bug Report and Feature Request Description Category Tool Experiments Experiment List New Exp2. CDCI AA 1 257 CDC polypeptide CDC2 AA 1 237 L CDC2 polypopside CDCI AA 1 297 CDC2 potypeptinde CDC2 AA 1 297 we i CDC2 potypepside CDCI AA 1 297 COC potypeptite COC AA 1 227 teres a COC polypeptide CDCI AA 1 297 a CDC polypeptide COC AA 1 227 L CDC polypapside CDC AA 1 297 1 CDC2 polypeptide CDC2 AA 1 227 a CDC2 polypapside CDC2 AA 1 297 1 CDC potypeptinde CDC2 AA 1 297 a CDC2 potypesside CDCI AA 1 297 a 9 E 10 12 ce Eaperimart Lat Logistic Score Sequest Fie Limit Mass Error pty Resswgn Collection kav n e rt as Pamias User Files Reduce FOR Limit Phosgho Reassign Subccilecnon _ esai n A Map Network Protocol View Lint Ascore Remove lrvaits Spec Find a Protein Q bertuan 20130 tins M 8 9 Kooere harga nata x kord Custore Load yr s ES PoP Excel asus mizera s 50 wd Bowe EEEE KID Category Tool Description Main Table Decoy DB Direction Describes whether peptide is a forwardor reverse hit in decoy database Peptide Sequence Displays the peptide sequence ofthe peptide record Phosphorylated sites are showninred Click to view the peptide spectra Phosphosite Displays the phosphorylatedamino acid s in the peptide Phosphosites inred denote pTyr containing sites Go To Subviews Export Filters Protein Name Additional Names Info Button Home Page Exp List Collection Sub Collection Experiment Load Custom Loa
3. 2009_Comparison4_042809 04 28 2009 1 single frac Label free Load Gokhan April2009 Comparison3 042809 04 28 2009 1 single frac Label free Load Gokhan_april2009_Comparison2_042709 04 27 2009 1 single frac Label free Load Gokhan April2009 ComparisonFr1 2 HybridSet 042409 04 24 2009 1 single frac Label free Load JurkatSip76042409_mascot_with3min 04 24 2009 2 single frac MS1 Labe Load _JurkatSip76042409_mascot 04 24 2009 2 single fraciMS1 Label Load Gokhan_April2009_RatLiverRefed_PHOSTIO_Fr22 04 23 2009 1 single frac Label free Load Gokhan_April2009_RatLiverRefed_PHOSTIO_Fr1 04 22 2009 1 single fraciLabel free HomePage Comparison JurkatSip76041709_mascot Q ix New Comparison i Browse 5 KG Comparison View This view is use to do any analysis on two state comparisons Each peptide record displays its name phosphorylated sites phosphoproteomic data in heatmaps display a free for all notes section and the peptide seguence Rollover the heatmaps for more detailed information on each datapoint Also clicking on the heatmap squares allows you to look at or validate the corresponding spectra The comparison view offers various statistical and biological filtering tools to help clean up the data set Peptides can be manual categorized into groups for further organization When the data set is set for publication several export options including pdf export for publication and raw data in excel format are available 00
4. PEPTIDE DEPOT SOFTWARE USER MANUAL Norris Hung Arthur Salomon Lab July 27 2009 Table of Contents 1 Introduction 2 Setting up Peptide Depot in Your Lab a Before You Begin b Installation guide c Accessing Peptide Depot 3 Navigating Peptide Depot a Homepage b Experiment List C Experiment View d Comparison List e Comparison View f Sample Queue Introduction to Peptide Depot What is Peptide Depot Peptide Depot is a custom FileMaker inteface that provides a central location for phosphoproteomic peptide data management and analysis Peptide Depot organizes peptide records by experiments and collections and provides an interface to easily browse through experiments This software also offers an interface to manage and analyze comparisons between datasets such as timecourses or wild type mutant comparisons Once viewing an experiment or comparison many tools such as spectra viewers protein network maps and condensed timecourse data heatmaps can greatly enhance phosphoproteomic analysis What You ll Learn This manual will show you how to guickly set up Peptide Depot in your lab and will guide you through all the functionalities available in different layouts of the software Setting Up Peptide Depot in Your Lab Install FileMaker server 9 or higher on your computer If you don t have a copy you need to purchase it from FileMaker corp Unzip the downloaded file to any folder After installation of FileMaker se
5. d DTA OUT Logistic Score Protein List Map Network User Files Protocols DTA OUT PDF Excel Raw File Seguest File Autofilter Limit Log Score Limit Mass Error Reduce FDR Limit Ascore Xcorr charge Limit FDR Redundinator Phosphorylated pTyr Limit Phospho Remove Invalid Spectra Mowse Score Forward DB only Displays the chosen protein name Hover over to choose other protein names Shows other possible protein names for the givenpeptide Click to show online database information onthe current peptide record e g HRPRD NCBI OMIM Pubmed Wikipedia Brings you to the home page Shows alist ofallexperiments withsummaries Use the drop down menu to pick a collection ofexperiments Use the drop down menu to pick asubcollection ofexperiments Use the drop down menu to pick anexperiment Loads the selectedexperiment Use this toload records from more than one experiment In order for this to work records need to be fromexperiments witha matching unique keywordin their names DTA raw MS MS Spectrafile OUT database search output mascot or sequest Probility score indicates the validity of peptide sequence identifications Shows all proteins captured by the dataset Move button to Comparison View Opens the user folder This view allows the user toview andedit protocols usedto create this dataset Exports the DTA OUT Exports aPDF ofall the records including heatmaps Exports all numerical and metadata as a
6. eriment Experiment Loader Comparisons Comparison List Experiment List This view lists all of your experiments and relevant summary information about each experiment To provide amore realistic picture of your experiment the summaries only take into account proteins that survive a 1 FDR threshold In addition you can arrange experiments by name date or collection by clicking on their respective labels at the top Tue 7 28 2009 Tue 7 28 2008 Tue 7 28 2009 Tue 7 28 2009 Tue 7 28 2009 Mon 7 27 2009 Mon 7 2712009 Mon 7 27 2009 Mon 7 27 2009 Mon 7 2712009 Mon 7 12712009 Mon 7 2712009 Mon 7 2712009 Mon 7 27 2009 HomePage Colection Q Sub Collection jurkat 2D1 3rd Experiment 75 woe Browse jurkat_2D1_3rd unassigned ERLIC ERLIC jurkat 2D1 3rd jurkat 2D1 3rd ERLIC YiYuan ERLIC 23min 072609 054447 ERLIC 22min 072609 032600 ERLIC 21min 072609 010717 BSAFTnrp2 400f 072609 022349 ERLIC 20min 072509 224820 ERLIC 19min 072509 202934 ERLIC 18min 072509 181042 BSAFThrp2 200f 072509 224358 BSAFThrp2 200f 072509 205409 ERLIC 7min 072409 195715 ERLIC 6min 072409 173818 ERLIC Smin 072209 073406 ERLIC 4min 072209 060124 ERLIC 3min 072209 042844 jurkat 2D1 3rd 1 Sm 01 071309 17215 Load Tells you where things are Easy 20 fiteres Unique OPOI Sites amg Experiment View This layout offers a view of all the pep
7. n excel database Opens the raw datafile for the entire experiment Opens a Zip file containing all database search output Combination ofmost commonly used filters limit mass error remove redundant kill IgG etc Many filters remove unnamed proteins Ig s that help reduce the FDR Custom Value Removes all redundant peptides Filters out all records with unphosphorylated peptides Filters out allrecords without phosphotyrosine Limits no phosphorylation sites per peptide Removes any manually invalidated spectra Customvalue Removes all records that are ahit against reverse sequences inthe decoy database Table rate Assigns current dataset toa particular species Used for protein interaction Comparison List This view lists all of your comparisons and relevant summary information about each experiment You can arrange experiments by name date or type by clicking on their respective labels at the top Comparison List Mion Name on Type ta Load _T03_3T3_SCX 07 20 2009 4 multi frac MS1 Label Load _T02_FOCUS_SCX 07 18 2009 3 multi frac Label free Load TO1 FOCUS IP 07 15 2009 1 single frac Label free Load Serio_rub1_tag 06 15 2009 1 single frac Label free Load BMMC timecourse 052209 05 22 2009 2 single frac MS1 Label Load Gokhan_May09_RLRefed_Fri_RepsComp_Hybrid 05 01 2009 1 single frac Label free Load Gokhan April2009 Comparison5 042809 04 28 2009 1 single frac Label free Load Gokhan_April
8. ng keywords Creates anew group fromthe selected peptides use the check box to select Exports the records in PDF format Exports all comparison data inan excel table Exports spectrain This area displays all currently applied filters Customvalue Customvalue Customvalue Filters out any peptide records that have below acertainamount oftimepoints Number of timepoints in manually set after clicking the button Filters out any peptide records that donot containaphospho tyrosine Displays the heatmap color key for current label free heatmap data Displays the heatmap color key for current SILACheatmap data Displays the color key for coefficient ofvariation percentage ofeach timepoint Displays number of proteins represented by the current dataset Displays the number of phosophorylation sites represented by the current dataset Displays the percentage of phosphorylation sites that are phospho tyrosine sites represented by the current dataset Displays the number of phosopho tyrosine sites represented by the current dataset pT Displays the number ofphosopho threonine sites representedby the current dataset ps Displays the number of phosopho serine sites represented by the current dataset Count Counts and refreshes the statistics basedonthe current dataset
9. o 2 pa Comparison View erres unstim 0 1 5 Peptide Depot PROTEOME Peptide Ore 10 unstim 0 1 3 5 10 san E ADAP a Betat integrin le Calmodulin a CBL B CBL a CBL a CD3 delta a CD3 delta a CDS delta B CD3 epsilon Y100 El 6 9 oe poreo e RVFDKDGNGY ISAAELR H sesave7a ag g _ Sena ola a K IKPSS SANAIY SLAAR P 5860 yera ag go D KIKPSSS ANAIY SLAAR P 8887 vera ag Oo eae KLIKPS SSANAIY SLAAR P vieo Zz 8 Eso i ioe R DDAQY SHLGGNWAR N vraa yeo Tn Dg Ea R NDOVY OPLRDRDDAOY SHLGGNWAR N Y188 Q g Functions o t JurkatSIp76041709 Load oO POF Excel Find a Peptido Make New Group Spectra by Counter twe Kech uF SLAC CV 0 A 12 Reduce FDR 3 IN Limit AScore a F3 22 j Limit Logistic Score 0 1 ta PPA Set Min Timepoints ai Grop Rebuild Cache Spectra by Name ze Count a 75 mada Browse Category Tool Description Main Table Grouping Check Box Use the check box to select proteins for anew group Protein Name Phosphosites Label Free Heatmaps SILAC Heatmaps Comments References Peptide Sequence Displays the default usually HPRD or manually choosen protein name for the peptide record Hover over toview other names Click to manually edit the name Displays the phosphorylated amino acid s inthe peptide An asterisk defines sites that are known inthe linked HPRD database Click to manually edit the phosphorylation site Dis
10. plays Label Free datain heatmap form Wild Type data by default Hover over to see more detailedinformation Goto Reading Heatmaps section for more information Displays SILACratio datain heatmap form Hover over to see more detailed information Goto Reading Heatmaps section for more information A free for all areatoinsert your ownnotes or references for each peptide Click to edit Displays the peptide sequence of the peptide record Phosphorylated sites are showninred Click toview the peptide spectra Go To Basic Functions Export SILAC Graph Filters Color Change Legends Statistics Home Page Comparison List Experiment Load Group Load Rebuild Cache Find a Peptide Make New Group Calculate Log Score Dist PDF Excel Spectra by Counter Spectra by Name Build ProteinNetVis Run ProteinNetVis Current DTA OUT Set All Raw Files SILAC Graph Applied Filters Reduce FDR Limit AScore Limit Logistic Score Set Min Timepoints Show pTyr containing LF SILAC CV Proteins Sites pTyr pY Brings you to the home page Shows alist ofall comparisons with summaries Loads the selectedexperiment Use the drop down menu to pick anexperiment Loads a specificgroup inthe experiment Use the drop down menu to pick a group Rebuilds the heatmap cache Use when heatmaps parameters have changed Caution this script may take up toa couple ofhours to complete Search for a peptide by usi
11. rver copy all database files fp7 in the FileMaker folder to FileMaker server s data folder PC Default c program files FileMaker server data Open FileMaker admin console and enable those files To verify FileMaker server and copied files open FileMaker pro 9 or higher if you don t have a copy you need to purchase it from FileMaker corp and in open remote menu connect to your FileMaker server s address and select to open peptide depot You shall be asked for username and password Use admin and default password brownuniv Since this username and password is used to open all related databases you may change password for all databases if you change one if server is OK you shall be able to open the database and explore data in FileMaker Go to the installer folder install MySQL and mysgl connector odbc Two types of MySQL database are provided in zip file under folder MySQL DB in sql format and folder dump Both contains three sample data set For sql you can execute in terminal via command mysql h hostname u username p lt your_path_to_the_sql sql For the folder type copy it to mysql s data folder and restart mysql server you may receive errors but it should work You will also need to setup a system DSN for mysql database Go to administrator tools and open data sources ODBC for management Goto system DSN panel and click Add New Choose MySQL ODBC 3 51 driver 3 51 14 as your driver
12. tides found in a experiment In addition to the peptide itself the software automatically determines a best fit protein for each peptide as shown in the protein name the associated protein can easily be changed by clicking on the protein name Details and statistics on each peptide are also displayed Filtering options are available to narrow down peptides based on statistical confidence and protein relevance In addition to the main experiment view there are a variety of subviews available that provide either amore technical or detailed view of each peptide or additional information on the experiment For example to learn more about a particular peptide you can click on the info button next to that peptide which will show you existing online knowledge on that particular protein When the experiment is ready for publication multiple export options are offered OOO Repti Depot PROTEOME 0 Experiment View 1 2 0 net ty et 3 m 4 CCOEACAGIGGEN 4 23 meta a Pept 3 bydreaytrystamine recestor _ B4 AD heat shock protein 4 xD heat shock protein vane apoptotic chromatin condensation CDC potypep re CDCZ AA 1 227 CDC polypasside CDCI AA 1 297 CDC polypep de CDC2 AA 1 2917 COC polypapsida CDCI AA 1 297 COC potypep rie CDC AA 1 297 POPU RRR R RRR RRR ERE RRR ERR RI CDC2 polypapside CDC AA 1 287 COC potypap de CDC2 AA 1 297 E mae CDC porypepside
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