KaPPA-View 4 Manual for Beginners
Contents
1. EI i 0 coo o 3 fF Ce a i i n m r EE ye PUREE TE p deea ig A a EEIE F TE LE i ni I j 1 i cggprert ceg uu E e FEITE f E E E F o UOT P Mai a s E oi et iTia H 3 Aro F M m Non metabolic genes which do not exist on the maps such as transcription factors can be analyzed By entering the gene IDs you can create simplified maps Hime Map Crear Hew Sore Wap Map Manner Myt Family AGRIS Myb Family AGRIS Gene List DBEBROBDOBOEBBOOBBRBRBODROORBROBRBROBDROOBDOBEGA BOBOOBBODOOOODOSBDRARABODRBBBOBOOBBOBBBBOOOHDADA BOOGOODODOOBOBOEBBHEOO In addition the users can also use the pathway maps prepared by themselves pa KaPPA View4 Manual for Beginners The GS GOGAT Cycle TEA Crete Up to four maps can be viewed at once in a single browser window a R pan asama 5 3 Osa ceeeeetoccenccacacccstereseseseeca Ea pa r a a 3 as taf Ht tt tte ta ci aoe Boge ee im ee es ae Ge C Pal The GSiA T Cee a agada roe me A Se e i m i ij lt rt Oe Ami a a y oo ae K Suie i et E te ack A ad wah ra p Saves 1 e O H aaam a a oa Tae amp T is 0 So eh oe eS T r i reic j E a 1 a d 2 D D a Furthermore gene to gene and or metabolite to metabolite relationships such as co expression correlations of ge2. Sly Data Then you get the comparative visualization of gene expressions and metabolite accumulations on a single pathway map m On Calvin cycle kinase 3 Phospho 3 Phoepho D glycaroyl Glycaraldehyde Tricsephosphate isomera D glycerate BAO a phcephate E ah S phosphate DE os Glycolate pathway A000 es Geos Dihydroxyacatone yp Glyceraldehyde 3 phoephate phoephate jehydrogenase ea NADP phoephorylating EA BE Ribulee bisphosphate m carboxylase small subunit Fructose bisphosphate aldolase M S EAR m5 Carton dixide Carbon dioxide T 2nSketolase D Ribulose Atmosphere 1 5 bisphosphate Alddase D Fructose 6 phoephate D Xylulose 5 phoephate D Fructose 1 6 m Phosphoribulckinase z bisphosphate D Ribulose 5 Fruct e bisphosphatase ho ephate D Erythroee 4 phosphate Aah DOEe 00880 on BREA isomera z x Fructoee bisphosphatase aldolase Glycearaldehyde 3 Alddase phcephate Sedcheptulose bisphosphatase ah EB Ah B D Ribose 5 phosphate oss DEE D Sedoheptulose 7 os E Uni00112 20080375 phcephate Transketolase D Sedcheptulose 1 7 bisphoephate 5 Other Functions The Simple Maps and The User Maps which are postulated to belong to a specific species are not utilized in the Universal Map Mode Representation of correlation lines is not available too 5
3. D Fructose 6 phoephate Aldolase Phogphcribulokinage D Fructose 1 6 5 phoephate bisphosphate ioe D Ribulose 5 phosphate Fructose bisphosphatase D Ribulose 5 phosphate 3 apimerase D Erythroee 4 phosphate To switch the Compared Experiment Pair click on Ath 2 The window will be redrawn with the data of Ath 2 and you will find Ath 2 is checked Compared Experiments C Ath 1 Ath 2 Ath 2 Transcript Sample Ath C Sample Ath D Show All Experiments Calvin cycle et Phosphoglycerate Gluconeogenesis kinase 3 Phoepho 3 Phcepho D glyceroyi Glyceraldehyde Triceaphosphate isomerase D glycerate 7 3 phosphate SEE g phoephate 3 phosp P oo Glycolate pathway O EDO a oT wd Glyceraldehyde 3 phoephate dehy jahydrogenase NADP phoephorylating Ribulcee bisphosphate Dihydroxyacetone phcephate carboxylase small subunit OH dioxide Transketolase Carbon dicxide Carbon D Ribulose Atmosphere 1 5 bisphosphate Phospheribulckinase 5 phoephate D Fructose 1 6 bisphosphate Fructoee bisphosphatase D Ribulose 5 phosphate S epimerase D Erythroee 4 phosphate Because metabolite experiments were not included in the Compared Experiment Pair Ath 2 the symbols for the metabolites O are all painted in white The detailed sample information is represente
4. 120 247 F Is Lele otk H L Fee PHARA qn P pm are ihri CMODIT 10 nd LSA OSN na a6 Mana NI iima Te oot UDI 100 04 Array Type Able Gi codes ath RAPS tiur RAPD tranecripis Aty Mice RAPS Wanscript Weal ase tne 3 Patua IJ Full lists of the probe IDs are also available from Download on the main menu appeared after logging in Select information as Data Type and find the Feature files The prefix of the file Ath_ Lja_ Osa_ and Sly_ stands for the species name as listed in the Statistics page For associating the gene expression values detected by probe on the microarray to the probe IDs on KaPPA View4 we prepared a Java tool KaPPA Average which is available from the following URL http kpv kazusa or jp kpv4 information tools_jp html On the preparation of metabolite data you have to know the compound IDs used in KaPPA View4 Please refer to a file named Uni_compoundInfo_yyyymmdd csv which is available from the Download menu or search the metabolite at Search on the main menu You have to input the experimental values as log scale for the probes negative to positive real values and as linear scale for the metabolites positive real values except zero Save the data files as text files formatted in comma separated vector CSV Se M KaPPA View4 Manual for Beginners 3 Data Analysis Basic Let s move on to data analyses on the pathway maps In this section we
5. KaPPA View4 Manual for Beginners 7 Inquiry Please inform us all of your questions and requests about KaPPA View4 if there are by e mail e mail kappa view at kazusa or jp please replace at to 8 Editing history 8 Editing Histories ver 1 2 September 21 2010 Some figures were updated 1 3 User Settings was updated Details of the operations were added to 5 1 Universal Map Mode and 5 2 Comparing two experiments in a species ver 1 1 July 1st 2010 The first release BA KaPPA View4 Manual for Beginners
6. pop up window A pull down list of Multiple Map will appear under the pathway map in the main window By selecting the combination name and clicking the Select button the pathway map area will be redraw to the Multiple Map Mode and the combination of the maps will appear BB KaPPA View4 Manual for Beginners Calvin cyde TCA cycle ieee eel Gereg Lh 7 ioe HW lsagdergipm oy Ea Tig ee bareen ou plein F mrak creche Tenens 8 Click Tiigri iE ai Ti g at Eug Pomno kiami sapa i sm i dli m biye iiien J And em pign de a r biie i n j 3 mi k form C E mbe pick E F chon piis Hive hia i bN FE pinampi cin T kban sapi im Le NFI Tu mimi om Aromatic amino acid biosynthesis ja ado Pana phoephete paihamy H F Ea mi m Hi v1 PL Tritt AA 1 eee Dairies Seite P i i Tpke ee z Aia rii geh m YP Dk mpi Ca eae ee Leh nee N HEF a a e aa h hdr Fy H mipainari dnain Miia amiin e Se a phere n elie I Oo oo LST AATA Multiple Map Multiple Map 1 w Select As described before the top left map of the Multiple Map is related to the pathway tree You can replace the top left map by selecting another map in the pathway tree To exit the Multiple Map mode select Single Map from the pull down list and click Select button Multiple Map Single Map PEUTEEITULELILILILELILELELLL ILE L
7. 2 Comparing two experiments in a species If there are several Compared Experiment Pairs for a species two of them can be represented simultaneously on a pathway map When several Compared Experiment Sets are set for one species a button titled Compare will be appeared Please check on two sets originated from one species and click the Compare button Ath Data3 show All Experime if Then you get the comparative visualization of two data 1 Set001 Transcript Ath A Ath B Metabolite Ath A Ath B 2 Set002 Transcript dark grown T678233 Metabolite Ath A comp test1 2 Calvin cycle Phosphoglycerate Gluconeogenesis kinase a DEAH 2 3 Phospho D glyceroyl oe Glyceraldehyde pyon hosphat phosphate 1 Glycolate pathway oon H U N Triosephosphate isomerase 2 2 3 phosphate gt OH ot 4 OS i Dihydroxyacetone Glyceraldehyde 3 phosphate dehydrogenas phosphate e NADP phosphorylating Ribulose bisphosphate carboxylase small subunit 8 Fructose BEG BED bisphosphate OB 2 BES BOB aldolase VOE BE 0 ma DE Carbon dioxide Carbon dioxide AES D Ribulose Atmosphere 1 5 bisphosphate Aldolase D Fructose 6 phosphate D Fructose 1 6 VY bisphosphate Phosphoribulokinase 5 phosphate ao 2 Ge Fructose bisphosphatase Dimm n mmA D Ribulose 5 phosphate 3 epimerase w
8. 36 KaPPA View4 Manual for Beginners Arabidopsis thaliana Myb Family AGRIS Arabidopsis thaliana metab Compared Experiments ICarbohydrate metabolisr H Amino acid nucleic acid ARI Seco lipids metabolism Show All Experiments Elsoprenoid metabolism Ath 1 Phenylpropanoid and shi Transcript Sample Ath A Sample Ath B a a id miiia Metabolite Sample Ath A Sample Ath B A Functional categories Simple Map Myb Family AGRIS E Mvb FamilwIAGRIS oR Boe EEEE iii iing ff hon felelel iinne oo Boe BOBS PFO E AEE A ROOOCOBBBBOBBeeeoeeeoeooos Coen oe oe Poe E A Be geno E Bon Belen fon ff Ben fon 0 0 Enig pe cE Eng E Boeken Palofefefafon Be oi x Create Simple Map 4 3 Creation of Multiple Map In KaPPA View4 at the maximum of 4 maps are simultaneously displayed on a single browser window This mode of view is called Multiple Map Mode which would help you to overview transcriptome and or metabolome changes in related metabolic pathways The user created Simple Maps 4 2 and the User Maps described later can be included in the Multiple Maps too Moreover the correlation lines see 4 1 are drawn across the each metabolic map Therefore the Multiple Map Mode provides an important basis for exploring regulatory mechanisms between genes and the metabolic pathways such as relationships between a transcription factor gene and metabolic pathways governed by the gene At the bottom of the pathway map you
9. Ath 1 A M Ath2 4G In the window you can choose which experiment in the pair is the denominator for the ratio calculation Analysis Compare Exp Name Exp Name Data Type Species Repetition Ath 1 Sample Ath A Transcript Arabidopsis thaliana 4 Sample Ath B Transcript Arabidopsis thaliana 4 Sample Ath A Sample Ath B Compare Exp Name Exp Name Data Type Species Repetition Ath 1 Sample Ath A Metabolite Arabidopsis thaliana 4 Sample Ath B Metabolite Arabidopsis thaliana 1 M2 Sample Ath A Sample Ath B Compare Exp Name Exp Name Data Type Species Repetition Ath 2 Sample Ath C Transcript Arabidopsis thaliana 1 M2 Sample Ath D Transcript Arabidopsis thaliana 1 M2 Sample Ath C Sample Ath D v Please choose one of two from the pull down lists 6 KaPPA View4 Manual for Beginners Compare Exp Name Exp Name Ath 1 Sample Ath A Sample Ath B Sample Ath A Sample Ath B gt When the experimental repetition was included in the data you can select here which repetition data should be taken account of the average calculations By checking off the repetition ID in the Repetition column the data is omitted for further analysis Repetition MiM In this tutorial it is not needed to change the settings Please push the Next button then the map browsing window will appear 2 Map View Map Wine 1 Select a name of species or Unmeisal from the pull down list 2 Click on a map ora
10. D Erythrose 4 phosphate QE m OB8BO PPE 0000 Fructose bisphosphatase aldolase Glyceraldehyde 3 Aldolase ye phosphate 5 3 Utilization from the outside systems KaPPA View4 serves public interfaces to utilize the system from the outside servers and application programs rr KaPPA View4 Manual for Beginners 9 3 1 URL link By describing URLs in the following formats developers can make links to the information page of the genes metabolites enzymatic reactions and pathway maps in KaPPA View4 After jumping the user will be recognized as a guest user and he she can continue to browse and analyze data with KaPPA View4 http kpv kazusa or p kpv4 geneInformation view action id At1 g58150 http kpv kazusa or p kpv4 compoundInformation view action id KPC00697 http kpv kazusa or p kpv4 enzymelInformation view action id R0000603 http kpv kazusa or p kpv4 mapView view action mapNumber 00006 The species can be specified too http kpv kazusa or jp kpv4 geneInformation view action id At12g58150 amp specie sName Arabidopsis thaliana You can check this action with a file ExampleUrlLink v html in the URL_link folder of sample data Please open the file with the Internet browser 5 3 2 POST Data Transferring Function This function is provided for developers of database sites and application programs where microarray and metabolome data are deposited In the usual way to represent the o
11. Pd etc Glutamate synthase o NADH typa 0 0 OO Ft type Pe O TA NH C L Giutamate Nitrate reductase NOs TCA Cycle Map Name Comment submit KaPPA View4 Manual for Beginners Input a map name in the Map Name field here input User Map 1 then click the Submit button The uploading will successfully finish with the following message The operation succeeded The uploaded User Map will appear in the pathway tree under the branch name User Map You can utilize it same as the default maps in your analyses _4 Data Analysis Advanced SS Arabidopsis thaliana gt User Map 1 Arabidopsis thaliana metab El Compared Experiments Carbohydrate metabolisr Amino acid nucleic acid sides L ipids metabolism soprenoid metabolism Ath 1 Phenylpropanoid and shi Transcript Sample Ath A Sample Ath B a knis aii Pee Metabolite Sample Ath A Sample Ath B E Myb Family AGRIS J Fa Glutamine synthetase Create Simple Map Ammonia Nitrite reductase HO N NH3 NO2 chtamine Nitrite a Nitrate reductase NO3 Nitrate 42 Show All Experiments The GS GOGAT Cycle Amino acids Nucleotides Chlorophyll a etc Glutamate synthase ri E NADH type fe o an ee A a HO OH O 2 Oxoglutarate bet TCA Cycle KaPPA View4 Manual for Beginners 5 Other Functions In this chapter we
12. Upload on the Main menu and select Correlation tab Experiment User Mag Correlation Type Gene Compound Conelation File 0 A Name Upload Here let s upload a sample data The sample data can be downloaded from the top page of the KaPPA View4 Press the Browse button and select a file named Correlation_Ath_Gene_v csv in the correlation folder in the sample data Input a short description of this file as Upload Test into the Name field The Comment field can be left as blank Press the Upload button After waiting for a while the uploading process will finish with the following message The operation succeeded The data uploaded here can be seen in the pull down menu of the correlation data Correlation Line Correlation Gene Upload Test No Lines Ath ATTED Ath ATTED Update Corr Ath ATTED Compound AthGeneCor_v3 1360 chips AthGeneCor_v3 1365 chips AthGeneCor_v3 1388 chips hormones 236 chips gt 0 81 tissues 297 chins gt 0 916 T ce 7 1 D ll stresses 298 chips gt E ee aie ree E hii Copyright 2004 2009 Kazu 4 1 6 Format of the Correlation Data Please open the file Correlation_Ath_Gene_v csv with Microsoft Excel 8B 4 Data Analysis Advanced As the row number in the file exceeds the Excel s capacity the whole data could not be shown But anyway you ca
13. ae serine Glycine and c PEPERIT NAPETE Aani S alyinisaii i ra ae 3 carboxy isocaproate oO o Histidine and nucleic B O f Branched chain aminc Miscellaneous amino tno E aa ads oO H Lipids metabolism a lt _ _ lt _ Oo oO p Leucine Valine and L Isolaucine hi UniG0008 2008 03 28 L Leuane ay Teens isoleucine metabolism Em E E E T N T F Then select the data as in the figure below and click the Update Correlation button OT 4 Data Analysis Advanced Correlation Line Correlation Gene Ath ATTED ll AthGeneCor v3 1388 chips gt 0 6 Compound Demo data from time course exps of drug treated Ath cells Update Correlation Smooth lines are appeared on the pathway map These lines indicate the relationships between the genes red and between the metabolites green Leucine valine isoleucine and alanine biosynthesi per Lysine threonine anc methionine biosynthesis m Glycolysis Katol acid raductoisomerase Acatolactate synthase Threonine dehydratase s 2 Oxobutancete 2 3 Dihydroxy isovalerate A Ber LN z caf z A i Dihydroxyacid A N i A ee K y V YA 2 Ketoisovalerate 2 isopropyimalate synthase Z W 3 Isopropyimalate dehydratase 2 D thrao Hydroxy 3 carboxy isocaproate Branched chain amino acid aminotrans
14. automatically log off when you close all the browser s o 7 2 Starting the Analysis windows too All your data uploaded according to the next section are going to be deleted from the system after log off 2 4 Uploading Experimental Data 2 4 1 Uploading of DNA microarray data Press the Temporary Upload on the main menu Main Temporary Upload Analysis Map View Search Download Temporary Upload Experiment User Map Correlation Experiment File CT 40a The following window will be displayed P Click the Browse button and select a data file As an example select a file named Sample_Ath_gene_v csv in the data_transcriptome folder in the sample data represents the version number Then click the Upload button Following window will be displayed to check the data Expanment eer Map Conelation Experiment File ELS Experiment Type Transenpt C Motabohte Paray Typi Sikl s Experiment Harme Repetition Humber Comment Sample Ath sample Ath E 1 1 Sample Ah C 1 2 Sample Ath O 1 2 Sample All Zero Conkrol 1 The file contains DNA microarray data of Arabidopsis Each of the data named Sample Ath A and P is from a single experiment C and D contain C KaPPA View4 Manual for Beginners experimental duplications In addition a dummy data Sample All Zero Control is included in the file As mentioned above this data is obtained fro
15. first show the procedure how to select the data from the uploaded data set and then explain basic functions for data browsing 3 1 Data selection for browsing In KaPPA View4 a unit of analyzing data is defined as a compared data between two experiments We refer this unit a Compared Experiment One Compared Experiment is comprised of a pair of gene expression data a pair of metabolite data or both of them Compared Experiment Gene expression data Exp Data 1 Metabolite data Exp Data 1 Let s try to make a Compared Experiment using the data uploaded in the section 2 Click Analysis on the main menu Main Temporary Upload Analysis Map View Search Download A data search window will appear Please check that Arabidopsis thaliana is selected as Species and TRANSCRIPT for Experiment Type Then press the Search button Tn 3 Data Analysis Basic Species Avabidopsis thaliana Selected Experiment T racreecrnpl Expenmnent Type TRANSCRIPT METABOLITE EP 5 Upload Usor All La Matahalaa Upload Date a E l Expenmant Sat Header AND COR Compared Expennnent Harm Data ial Sathi Expennent Date Hiad r z ax z CAND COR Aaa caran Search Reset On the lower part of the window a list of data which are currently available will be displayed Showing 10 7 per page Showing 1 2 of 2 Set ID Set Name Array Type No of Exp Uploaded Date Rel
16. map stegory on the metabolic pathway ree 3 2 Data browsing on the maps Using the data registered in the previous section let s browse the data First of all please select Arabidopsis thaliana from the pull down list on the top left T 3 Data Analysis Basic Carbohydrate metabolisr Amino acid nucleic acid Arabidopsis thaliana 1 Select a name of species or Universal from the pt p H Lipids metabolism i l i Universal i soprenoid metabolism 2 Click on a map or a map category on the metabolic 5 Arabido ae thaliana Phenylpropanoid and shi Lotus japonicus H Gene families and misce Functional categories Oryza sativa Solanum lycopersicum 1 ipids metabolism Hennrannid matahnaliem Next select one of pathway maps in the metabolic pathway tree An example selecting Calvin cycle is shown here Symbols on the pathway maps are painted in color according to the data Arabidopsis thaliana Iv Calvin cycle Arabidopsis thaliana metabl amp ElCarbohydrate metabolisr Toe et ai CO fixation and cen M Athi C Ath2 Calvin cycle Show All Experiments Compare lGlycolate pathw Ath 1 Glycolysis glucd Transcript Sample Ath A Sample Ath B i Phosphoenolpyr Metabolite Sample Ath A Sample Ath B ETCA cycle E Glyoxylate cycle ae Calvin cycle Glycerol metabc linasa a aes 3 Phoepho Ph ho D gl cl Ideh 2 Triceep osp isomerase a Mono di and oligos lt D g
17. metabolite to metabolite co accumulation data calculated from a series of metabolomics data obtained from a drug treated Arabidopsis cultured cells in our laboratory is also available as a demonstration data Let s try the operation As an example please select a metabolic map Leucine valine isoleucine and alanine biosynthesis of Arabidopsis thaliana Arabidopsis thaliana pas i Leucine valine isoleucine and alanine biosynthesis a S e a EE Lysine threonine anc El Arabidopsis thaliana metab _ Glycolysis methionine biosynthesis 7 A 7 Katol acid reduct Acatolacate synthase Threonine dehydratase Carbohydrate metabolisn Serene tes oe Ree ene 2 3 Dihydroxy isovalerate L Threonine El Amino acid nucleic acid o Dihydroxyacid dehydratase 2 Acatolactate El Aspartate and related ElGlutamate and relatec A l y Branched chain amino cetolactate synthase acid aminotransferase Alanine aminotransferase BOGOR ine ElGlucosinolate metabo 2 Aceto 2 hydroxybutyrate F i 2 Ketoisovalerate L Alanine Leucine valine isoleu Katol acid raductoisomerase 2 lsopropyimalate Leucine valine ieee 2 3 Dihydroxy 3 methylvalerate Leucine valine z 3 Carboxy 3 hydroxyisocaproate i N i i i 3 Isopropyimalate Dihydroxyacid dehydratase Aromatic amino acid i dehydratase 3 lsopropyimalate dehydrogenase TATE eg
18. of such approaches and it can list up co expressing genes of Arabidopsis for a query gene of researcher s interest KaPPA View4 provides a function to overlay gene co expression data onto the pathway maps Data representation in this manner helps to grasp the gene to gene relationships on the aspect of metabolisms KaPPA View4 can represent metabolite to metabolite correlations too As an index of co expression between the genes correlation coefficients have been typically used Hence the functions of KaPPA View4 concerning to the co expression of the genes or co accumulation of the metabolites are referred like correlation functions However the data which KaPPA View4 accepts is not restricted in the correlations Any data which represents gene to gene or metabolite to metabolite relationships as numerical values can be utilized For example protein to protein interaction data described by O or 1 is acceptable Please try to project your own ideas onto the pathway maps with KaPPA View4 26 KaPPA View4 Manual for Beginners 4 1 1 Viewing the Correlation Data The following control panel is displayed under the pathway maps Here the users can choose the correlation data to view Correlation Line Correlation Color Range Number Gene No Lines RED gt fo Compound _ No Lines gt E foc fho Hino so Update Correlation In the defaults several gene co expression data provided by ATTED II can be selected One
19. valine isoleucine and alanine biosynthesis __ Lysine threonine anc Glycolysis methionine biosynthesis lt lt a Ketol acid reductoisomerase Acatolactate synthase Threonine dehydratase L Threonine 2 3 Dihydroxy isovalerate 2 Acatolactate Branched chain amino A acid aminotransferase Dihydroxyacid dehydratase Acatolactate synthase Alanine aminotransferase 2 Aceto 2 hydroxybutyrate ERDROEN Wels 2 Ketoisovalerate Katol acid reductoisomerase 2 lsopropyimalate synthase 2 3 Dihydroxy 3 methylvalerate Dihydroxyacid dehydratase o 3 Isopropylmalate dehydratase E 2 D threo Hydroxy 2 Kato 3 methylvalerate S carboxy isocaproate Branchec chain aminc acid aminotransferase Branched chain amino acid aminotransferase aaa j Vali E Leucine Valine and anes L Isolaucine isoleucine metabolism Uni00008 2008 03 26 L Leucne In this figure three red lines are drawn However three gene symbols on the left hand side are for the same gene As shown here by setting a number in the Number field users can filter the correlation data to display the lines of highest values in the restricted range When you would like to view the correlation lines of the lowest values in the range select Low from the pull down list Range Number afi v3 High 0 9 1 0 0 6 4 1 3 Details of the Correlation Data Displayed
20. 0 6922613 0 11 Ati gO1090 1 487257461 0322357127 0615823494 0518919149 0 99989596 0 99000439 0 12 Atig01100 1 285759095 00542067 0 33525193 0 39955302 O0 756790904 0 754720246 0 13 Atig01110 057385919 0383093021 1 061435081 1 24052402 0 6461393 055626915 0 KaPPA View4 Manual for Beginners A z T D z F G 7 1 compexm Sample Ath A Sample Ath B Sample Ath B Sample Ath Sample Ath C Sample All1 Control 2 rep 1 1 2 1 2 1 3 KPCOO001 657 124 125 5 29839 5 34677 1 4 KPCOO002 9 9967 10274 0 0009 0 0009 1 5 KPCOOO003 241164 664 664 279 14815 275 60648 1 6 KPCOO004 122 18 18 6 77776 6 66667 1 7 gt KPCOO005 5140 aie 377 13 8172 13 51075 1 8 KPCOOO06 463 D1 51 9 07843 9 39216 1 9 KPCOOOO 458 4281 22 438831 0 001 07 0 001 OF 1 10 KPCOO008 504864 752 772 67 13298 66 69149 1 11 KPCOO003 53 235794 230191 0 00022 0 00022 1 12 KPCO001 0 64323 890 696 72 27303 72 54719 1 13 KPCO0011 91 1299 1294 0 07005 0 07159 1 Fig 2 5 2 Sample_Ath_met_v csv You can see two rows of header in both the microarray and the metabolite data files The first row starts with arrayexp or compexp In the uploading process the KaPPA View system recognizes the experiment type by this cell The subsequent columns of the first row are the data names The second row always starts with rep and the subsequent columns represent the repetition numbers Remember that the Sample Ath C and D of microarray data contained experimental duplic
21. 2 hydroxybutyrate Katol acid raductoisomerase 2 lsopropyimalate synthase 2 3 Dihydroxy 3 methylvalerate 3 lsopropyimalate Dihydroxyacid dehydratase dehydratase 3 Isopropyimalatp dehydrogenase 2 Keto 3 methylvalerate 3 carboxy isocaproate Branched chain aminc Branchec chain amino acid aminotransferase acid aminotransferase Leucine Valine and isoleucine metabolism gt rs lt r rr L lsoleucine Uni00o008 2008 03 28 L Laucine 4 1 3 Filtering the Data to View Number of the Lines When the ranges are changed by the way described above a text like 3 is displayed in the Number column It was 29 before the filtering Number High 0 6 The value after the slash means the total number of the correlation lines currently represented on the maps The redundant relationships of the same combination of the genes are excluded from the count As there are in the case that the same gene is drawn at multiple places on a map the indicated number could be different from the line numbers drawn on the maps Correlations calculated in an element self correlations are excluded from the count even if they are included in the uploaded files ns 22 4 Data Analysis Advanced Anyway please enter 1 in the field written as 0 see below and click Update Correlation button Number Then only the highest correlations is drawn like the following figure Leucine
22. 2780 E 0 9159 0 148 0 7678 1 467 0 978 0 489 0 000 0 489 0 978 1 467 Ath 1 Ath 2 gt linear log Ratio v show symbol 3 3 The Bird s Eye Map We described so far about the pathway maps placed at the lowest tier leaf of the Pathway Tree Now please click the middle tier of the tree branch Following map will appear 3 po KaPPA View4 Manual for Beginners Arabidopsis thaliana zl Amino acid nucleic acid and nitrogen containing derivative metabolism ElArabidopsis thaliana metab View Thumb Nails View Birds Eye Map i bolisn ElAmino acid nucleic acid Name Y Select Display Mode Aspartate and related Glutamate and relatec T Bacao sane Eon Amino acid nucleic acid and nitrogen containing Glucosinolate metabo derivative metabolism Aromatic amino acid 1 i Aspartate and related amino acid metabolism j si Serine Glycine and c Aromatic amino acid metabolism Histidine and nucleic Miscellaneous amino Lipids metabolism lsoprenoid metabolism Phenylpropanoid and shi Gene families and misce Kil gt These maps are called Bird s Eye map in KaPPA View4 Each of all the pathway maps included in the branch is represented as bar indicator bar At the first time to display Bird s Eye map the names of the pathways are displayed in the bars You can change the contents in the bars by choosing the items fr
23. 4 t 04 77 mor C i oo T 143 Miscellaneous amino acid related metabolism 7107 EM 7610 T 05 C7 M 02 T 08 C M 08 G 07 CJ mors CD L Gono Imon aa Se 7 26 E os M T u31 E M ete The numbers written beside T or M numerators show the numbers of the correlation lines on the pathway maps The numbers after the slashes denominators indicate the numbers of the genes or metabolites drawn on the maps For the indicators of the middle tiers both of the numerators and the denominators are the sum of the line and element numbers included under the tier The color of the bar is decided as follows When defined D logio line number element number of a map Dmax The maximum value of D among the maps under the current tier and Dmin The minimum value of D among the maps under the current tier the bar of the map having Dmax is painted in the strongest color red and the map of Dmin is painted in the weakest color green Therefore the maps having dense relationships are painted stronger colors Correlations calculated in an element self correlations are excluded from the line number counting even if they are included in the uploaded files 4 1 5 Uploading Users Own Correlation Data Users can upload their own correlation data to KaPPA View4 and utilize them in the analysis 32 KaPPA View4 Manual for Beginners Click Temporary
24. DA BHEOUOBEBEBEBEBBOBRAEBE B BA KaPPA View4 Manual for Beginners A full list of gene IDs used in the KaPPA View system is available from the Download page v A Calvin cycle El Arabidopsis thaliana metab Click Create Simple Map button appeared under the pathway map tree Arabidopsis thaliana Carbohydrate metabolisr ECO fixation and cent Calvin cycle lt Phosphoglycerate Gluconeogenesis s kinase Gin cycle 3 Phoepho D glycaroy Glycaraldahyde Triceephosphate isomerase g ycerate hoephat 3 phosphate P phcephate Glycolate pathw Glycolate pathway e ass m m ma O O C1 Dihydroxyacatone f yp Glyceraldehyde 3 phoephate phosphate Glycolysis glucc dehydrogenase NADP phoephorylating B Phosphoenolpyr Ribuloge bisphosphate f A TCA cycle carboxylase small subunit Chreavis mr Glyoxylate c ycle bisphosphate Glycerol metabc D Ribulose Atmosphere 1 5 bisphosphate Mo Polysaccharide metat Aldolase no di and oligos E Phosphoribulokinase Miscellaneous carboh D Fructose 1 6 bisphosphate Fructoee bisphosphatase D Ribulose 5 Amino acid nucleic acidy phosphate D Ribulose 5 phosphate D Erythroee 4 phosphate lt i gt 3 apimerase Create Simple Map 0o n00 m a a Ribose 5 phosphate is
25. Guest Login Password in Click Create Account on the top right of the browser window Online Help Create Account Log off GUEST Temporary Upload Analysis Map View Search Download A pop up window will open Enter your account name and e mail address and then press the Submit button co KaPPA View4 Manual for Beginners Create New Account Login Name Submit An e mail will be sent to you immediately and it informs you the access password 6 2 Expiration of the Power User The Power User account will be automatically deleted in 30 days after latest logging in All the data uploaded by the Power User will be deleted too An alert e mail will be sent to you in 21 days after last login To keep your account please login again before the expiration date 6 3 Power User Login Enter your name and password in the fields on the top page and then press the Login button KaPPA View 4 Kazusa Plant Pathway Viewer Home Overview News Statistics Download Link Publication Contributor AboutUs Login Welcome to KaPPA View4 Classic Go to My Page Name username Enter Password ececceccce Please click the button to start All the data is freely accessible for publi Login users More advanced you can save your own data on KaPPA View4 by creating your account Enter and click Create Account for an immediats registration free Gene information for Oryza s
26. KaPPA View 4 Kazusa Plant Pathway Viewer KaPPA View 4 The Kazusa Plant Pathway Viewer Version 4 0 Manual for Beginners ver 1 2 ee Kazusa DNA Research Institute Table of Contents Table of Contents PMs MINEO CUCU Oh eere a a E E EE E EEEE ENE 1 1 1 Overview of KaPPA VIew4 ccccccccssssscceeeeeecseesseeeeeeeeseneaaseeeeeeseeessaaseeeeeeesssaaagseeeeeeeess 1 1 2 Downloading Sample Data ccccccccccccccccceeesseseeeeeeeeeesaeeeeseeeeeeeeesseseeeeeeeeessaaageeeeeeeees 4 Te UST oe ee EEE E E EEE E EAEE 4 1 4 Other Manuals cccssessesseesseeeeeeeeeeeeeeeeaaaeesssseeeeseeeeeeeeeeeeeeaaaaaaeasaeeeeeeseeeeeeseeeeeensaaaaas 5 2 Starting the Analysis Login and Upload of Experimental Data 6 A EOI ee E E E E E 6 2 2 The Main INIGT NU iccseecncasaamancnsnnanaencnnvetieeshesesdannsidsariaanonscanvataspeaanwaieaneoeinuieausedeiaatteeskauetatactns 6 BOO acto E teers ces oe E E P E A E E E A E E toeeate eee ees 7 2 4 Uploading Experimental Data cccccccccccseseeeeceseeeeeeseeeeseeeeeeeaeeeseseeeeeseaseeesseeeesnageess 8 2 4 1 Uploading of DNA Microarray data cccccccccceseceeeeeeeeeeeceeeeeeeeesueeeeeessaeeeeessaaaeeeeessaeeeeeesaeees 8 2 4 2 Uploading of Metabolite Data cc eeccccccsssceccecssseeeeeeseeeeeeeeeeeeeeeeseeeeeeeseaeeeeeeseaeeeeeesseeeeesessaess 9 2 5 File Format of the Experiment Data cccccccccsssecceeeeeeeeeeeseeeseeseeessaeeeseeegeesaaeeee
27. LL ELE LLL ELL LLL ELLE LLL RR ee rr 4 4 Utilization of User Maps o 3 4 Data Analysis Advanced Users can create their own pathway maps and utilize them in KaPPA View4 It would help you to analyze non metabolic genes which don t exist on the default maps to make more beautiful pathway representations to analyze with the maps with careful curation of the gene assignments and so on The user created maps User Maps have to be prepared in Scalable Vector Graphics SVG format We recommend doing it with a freeware Inkscape Please refer to the Manual of User Map Creation for the details User defined gene IDs and compound IDs could be included in the User Map data If the IDs and their values are written in the analyzing data the symbols of the user defined elements are painted with color like as the pre drawn ones on the default maps Let s look at the way to upload and utilize the User Maps here with a sample SVG file Click Temporary Upload on the Main Menu and select the User Map tab Experiment User Map fCorrelation User Map File ia Upload Click the Browse button and select a file named UserMap_GS GOGAT_v sve in the userMap folder in the sample data Press the Upload button then a preview will appear Temporary Upload Experiment User Map Correlation User Map File AR Upload Map Preview The GS GOGAT Cycle Amino acids Nucleotides Chlorophyll
28. You can see what genes and metabolites are connected by lines in the current map and what the correlation values are by clicking the Correlation List BO KaPPA View4 Manual for Beginners Element List Correlation Li thy Histogram Color Legend Download Plain Map Print Map Alternatively when the mouse cursor is over the correlation lines the lines are highlighted and the gene IDs and the correlation values are shown in a tool tip 2 0 xobutanoate L Thraonina 4 1 4 Displaying densities of the Correlation Lines on the Bird s Eye maps When the Correlation is selected for Display Mode in the Bird s Eye Map users can see the line numbers on the pathway maps Display Mode REMUE In the similar way described above please select the data to view and the filter conditions at the bottom panel 4 Data Analysis Advanced Amino acid nucleic acid and nitrogen containing derivative metabolism Histidine and nucleic acid metabolism ra Aromatic amino acid metabelism T 5101 LL mM ort Cd Aspartate and relsted amino acid metabolism 263 a i 27 7 T a MaA 1407 E 175 E ao Go oe EUO lr oen nana oe cen 1 013 CA mow Coy 7 157 EE 0 121 a THe aay M a2 N 7 07 C m 0 5 M 0 23 7 16 B i 012 18 Ly M 010 7 17 GE saw 26 lL T 01 C M07 Cc O02 Gluccsinolate metabolism 7214 Sy M 415
29. ated Data gt KESTI pn ee accede 8 2009 10 28 Data gt TempSet_000001 TempSet_000001 AGI codes 4 2009 12 09 The data uploaded in the section 2 1s registered under the Set Name Tempset_000001 A list of experiment data contained in the experiment set is shown by clicking the arrow head P presses iY TempSet_000001 TempSet_000001 AGI codes 5 2010 01 03 Exp ID Exp Name Comment Type Cd TempExp_000001 Sample Ath A quantitative Cd TempExp_000002 Sample Ath B quantitative Cd TempExp_000003 Sample Ath C quantitative Cd TempExp 000004 Sample Ath D quantitative Cd TempExp_000005 Sample All Zero Control quantitative Click the data icon E3 on the left of the experiment data Sample Ath A The name is appeared in the top right panel Selected Experiment Transcript Sample Ath A Metabolite Compared Experiment Name Set001 Add Clear All JA Oo 4 Then click on the data icon of Sample Ath B The experiment name is appeared as the second one Ero T KaPPA View4 Manual for Beginners Selected Experiment Transcript Sample Ath A Cd Sample Ath B Cd Metabolite Cd Cd Compared Experiment Name Set001 Add Clear Al Let s try to select metabolite data In this state please check on METABOLITE for experiment type and press Search button V TempSet_000002 TempSet_000002 4 2010 01 03 Exp ID Exp Name Comment Type Cd TempExp_000001 Sample Ath A quantitativ
30. ation Therefore the repetition numbers of it were set to 1 and 2 see the second row of Column D and E in the figure 2 5 1 The experimental values see below of the repetitions within an experiment are averaged for each gene or metabolite and the representative values are utilized to display on the pathway maps The third and the subsequent rows describe the experimental data The first column is IDs for microarray probes microarray data or for compounds metabolite data and the second to the last columns are the experimental values The list of the valid probe IDs that KaPPA View can accept are written in the statistics page which is linked from the top page of the KaPPA View site T 2 Starting the Analysis KaPPA View 4 Kazusa Plant Pathway Viewer Statistics Download Link CAT Home Overview News Login Welcome to KaPPA View4 Classi Enter Gene and Probe infromation Spe Geena I Arabidopsis ihaliana AGI codes tram TARH Ona sativa RAP O8 build 5 1oou ryza saiva RAP DE transcnpl HAF DE build i fanecipis Tomate GeneChip proves SAI Iranersicum Tomato Ameti Loti ppanc Mipakoyusal jp arimotation fishin ine mar Amaman Shee an Pree Rin AARSE Publica Go tc Narr Prai 1 Sampii same a6 ene Ar gTzdan Same as ene i Os0ighi32832 AAP DB bud transeripls I Pe an arr OADW Aanoetric Rice GeneChip probe ID Os 6 1 1 31_6_al same a6 ane Same as Gene Les
31. ativa was changed August 15 2010 The transcript IDs from RAP DB have been used for gene IDs of Oryza sativa We changed the gene IDs of it to the locus IDs from RAP DB so that the correlation data of rice could be represented The old settings are preserved as the species name Oryza sativa RAP DB transcript so your microarray data described as the transcript IDs can be analyzed with it The data also can be analyzed in the updated Oryza sativa by selecting the array type RAP DB transcript A main window for Power Users will be displayed T 6 User Account Pere 1 3 Mair The Side Menu for Power Users feid of the Side Mer Menu Bar Tempolary Upload 6 4 Power User s Menu There is a Power User s Menu Side Menu on the main page Personal gt 5 Fa Ei Experiment Upload User Map Upload Correlation Upload Personal Data List Utilities ri gt Password Change Profile Edit Personal Block Here Power Users can upload their own experiment data User Map data and correlation data The uploaded data will be stored in the KaPPA View server and kept until the expiration of the account The data is strictly administrated in the system and is never seen by the other users Management of the data i e deletion and edition is available in the Personal Data List Utilities Block Password changing and editing of user information is available here s n
32. briefly introduce other functions of KaPPA View4 Please refer to the Manual for Users for the details of each 5 1 Universal Map Mode There is an item named Universal in the pull down list for species selection Arabidopsis thaliana Universal i Arabidopsis thaliana Lotus japonicus Oryza sativa Solanum lycopersicum When the Universal is selected Universal Map Mode the information of all the species is displayed on the pathway maps You can see the differences of the gene assignment to the enzyme reactions between the species Citrate synthase Sy Aconitase hydratase OOOO sy O sly OO osa OO oss Oo0000 ah Citrate OOO ath Gs Aconitate Aconitase hydratase o HHo z a beta Oxidation of saturated fatty acid COA Glyoxylate e _ isocitrate lyase eeeeces Succinate When there was not an enough space on the map to represent all the gene assignment a box written as will appear Clicking on the box a pop up box is displayed to view the all ns Te 5 Other Functions Citrate synthase Acalyl CuA mi Aconitasa hydratase ath O Lja O os OOO cis Aconitate sy O isocitreie lyase The species displayed on the pathway maps could be changed by the user by clicking the Select Species button at the bottom of the map Ribose 5 phosphate isomerase Glyceraldeh phoephate D Ribose 5 phosphate o D 3 Uni00112 20080375 pho Tran
33. can see a button named Add Related Map Click the button then a pop up window will appear Sedcheptulose bisphosphatase O D Ribose 5 phosphate a D Sedoheptulose 7 Unid0112 2008 03745 phoephate Transketolase When Universal is selected the button is not displayed y 4 Data Analysis Advanced In the pop up window you can set a combination of the maps In the Multiple Map Mode top left panel of the tiled maps is automatically set to the one which is currently viewed selected in the pathway tree of the main window Therefore Current Map is written in the top left panel and you would select here the other 3 maps 1 e top right bottom left and bottom right panels By clicking a map name in the pathway tree of the pop up window thumbnail of the map will be added to the preview area sequentially in the order above Plant metabolic pathways Multiple Map Create Carbohydrate metaboli Amino acid nucleic ac ipids metabolism soprenoid metabolism Phenylpropanoid and s Gene families and mis Functional categories User Map Current Map E My Map 1 A v Clear Clear You don t have to select all of 3 maps Selection of only one or two maps is acceptable After selecting the maps enter a name of the combination in the Name field and click the Add button Name Multiple map1 e Next click the Redraw button to refresh the main window After that close the
34. compound on the metabolic pathway maps By default approx 150 maps are available corresponding to Arabidopsis rice Lotus japonicus and tomato Upload TAT wie sg m KaPPA View 4 Kazusa Plant Pathway Viewer Display on the browser KaPPA View4 Server User s PC On the pathway maps genes and compounds are represented by squares and circles respectively The symbols are painted in different colors depending on the values such as changes in the ratio between two experiments and the amounts detected in one experiment ee Ge 1 Introduction Aromatic amino acid biosynthesis m Ratio Expression an mica pha pIE i M em rarer fo n Exp A Exp B Accumulation feemmes ain oye i ae amount Color Eei High Large ora a a a O O aunt Tea LO O P a I U Dope ee eee E Tar L No change i oe i i Ain bacmeyreteeaza E 7 g DEN banyan thera rivai p arr bas pearl oral ye ee ae a g a Small On the Bird s Eye View Maps you can view the summarized values for all maps and find out the pathways which have changed considerably i i LL Sa T ns bh es Se 7S e oo ake Sie E a io G re a en El 1 7 ae ae E WE G ra rae jo e e Sr a e M a M Cee i a a A Ses Fo mam CEN ii La 1 st a ees U m Ce uum ures maru xe pe 7 pU o o o G Cte a es CS l a eee ee o ee DEII i _
35. d Experiment Type Transcript Metabolite Select Select lt lt lt Arabidopsis thaliana WLU AU z Solanum lycopersicum The Experiment Type is automatically set to Metabolite by the Species Experiment Namg auto recognition of the file format Then press the Submit button Uploading will be finish with the following message The operation succeeded 2 5 File Format of the Experiment Data Here we show the file format of the experiment data Please try to open the files uploaded in the section 2 4 with Microsoft Excel Fig 2 5 1 Sample _Ath_ WO gene_v csv A B C D E F G H I 1 arrayexp Sample Ath A Sample Ath B Sample AthC Sample AthC Sample Ath D Sample Ath D Sample All Zero Control 2 rep 1 1 1 2 1 2 1 3 At1g01010 0 82847516 0646901298 1190483986 1 07724172 1 15233451 1 010858621 0 4 Atig01020 1 52244998 052435562 0 16694342 0 22203733 1 269603413 1 352828554 0 5 Att gO1030 11188739492 1 41118869 1 26662646 1 29635268 0 01673949 0 06297865 0 6 Atigo1040 1 21811202 0 96145002 0 07469906 0113924618 1 0868689507 1 064449577 0 7 Att g01050 0 646493581 0126151787 1175051608 1170206475 O529977397 040708899 0 8 Ati g01060 0 809261597 1566740565 O579037425 oO609907787 1 405337858 1 331461944 0 9 Atigo107O 1 204413521 0 88953735 061000164 0611701704 1 334508091 1 561604352 0 10 Ati gO108O 0904213049 1 059304533 0077268201 0060123451 O062478865
36. d above the pathway map As exemplified here several Compared Experiment Pairs can be set in a single analysis and users can browse each of them by switching the data It is suitable for browsing related data sets such as time course experiments 3 2 4 Details of genes metabolites and enzyme reactions You can get the detailed information about the symbols for genes metabolites and enzyme reactions by clicking them on the pathway maps A window will ns a 3 Data Analysis Basic open and you can see descriptions of the element values of the experiments and so on Gene Information At3g12780 Gene ID 14 TAIR Annotation 1 AT3G12780 1 PGK1 PHOSPHOGLYCERATE KINASE 1 phosphoglycerate kinase 1 AT3G12780 1 similar to PGK PHOSPHOGLYCERATE KINASE Arabidopsis thaliana TAIR AT1G79550 2 similar to phosphoglycerate kinase putative Arabidopsis thaliana TAIR AT1G56190 1 similar to chloroplast phosphoglycerate kinase Populus nigra GB BAA33803 1 Description similar to Phosphoglycerate kinase Medicago truncatula GB ABE80998 1 similar to Phosphoglycerate kinase chloroplast precursor GB P50318 contains InterPro domain Phosphoglycerate kinase InterPro IPR001576 Currator summary nuclear phosphoglycerate kinase PGK1 Arabidopsis thaliana Map 1 Calvin cycle 2 Glycolysis gluconeogenesis 1 R0011107 Bani 2 R0011202 Experiment Value ID Color Ratio Sample Ath C Sample Ath D At3g1
37. e Cd TempExp_000002 Sample Ath B quantitative Cd TempExp_ 000003 Sample Ath C quantitative Cd TempExp_000004 Sample All 1 Control quantitative The second data temporary uploaded is registered under the Set Name TempSet_000002 Expand the list by clicking arrow head P and select experiments Sample Ath A and Sample Ath B The top right panel will be like this Selected Experiment Transcript Sample Ath A Sample Ath B Metabolite Sample Ath A Sample Ath B Compared Experiment Name Set001 Add Clear All Jo a a Let s register this combination of experiment data as a Compared Experiment Please type Ath 1 in the Compared Experiment Name field and press Add button At the bottom of the top right panel the Compared Experiment Name will appear Compared Experiment List W Ath1 4G Next gt gt ns a 3 Data Analysis Basic You can register more than one Compared Experiments Try to register next One Select TRANSCRIPT for experiment type press Search button to show the data list select the experiment data Sample Ath C and Sample Ath D and register this setting as Compared Experiment named Ath 2 Selected Experiment Transcript Sample Ath C Cd Sample Ath D Cd Metabolite As described here you can register a Compared Experiment Pair which has only microarray data Please push Next button to go to the next window Compared Experiment List M
38. eaaases 46 5 3 Utilization from the outside SYSTEMS ccccceeccccseeeceeeceeeeeceeeeceeeeeeeeeseeeeesaueeeessaaeeessaees 46 ra URLAR eee en ee ee ee eee ee enn nn ee eee eee ee 47 5 3 2 POST Data Transferring FUNCTtION ccccccccesseeceeceeseeceeceeseeceeeaeeeeceeeeeaeeceeseeeeceeeeeaeeeeeeeas 47 Os USEr ACCOUNTS vac wis tess rionn E TE E Re 50 6 1 Creating an ACCOUNT setrane E A EREE 50 6 2 Expiration of the Power USE cccccceccssseseeeecaeeeeceeeceeeeeeeeeaeeeeeeeeesaeseeeessaaeeeeeenaaees 51 6 3 Power User LOGIN serasroera sinr 51 6 4 Power User s MONG x casceacoccesedeascesencsie5ec asaveeseusounssedtanaedennaessendesvaedsaderecsudsnsasieasaossaedeoess 52 eNO UY aces acess cee ceaatue saveutapcanareasautes EE 53 os CEN a SOUS S sree cesses eters EE E E 54 Table of Contents KaPPA View4 Manual for Beginners 1 Introduction KaPPA View4 http kpv kazusa or jp kpv4 is a metabolic pathway database which is aimed to better understand metabolic regulation and to generate hypotheses from huge public available omics data 1 e transcriptome metabolome and co expressions of the genes This manual guides major functions of KaPPA View4 to facilitate the beginner users to understand how to use the system 1 1 Overview of KaPPA View4 When you upload your own DNA microarray data and or metabolite data to the system through the web browser KaPPA View4 displays the fitting of data for each gene or
39. eeeeeeessaaeeeeeeeeesaaaaseess 26 4 1 1 Viewing the Correlation Data ccccccccsceeecccceeeeeeeeeeeeeeeeeeeeeeeeceeeeesseeeeeeeeeessesaaaeeeeeeessaaageess 27 4 1 2 Filtering the Data to view Range of the Correlation Values cceeeceeeeeeeeeeeeeeeeeeeeeeeeeaeees 28 4 1 3 Filtering the Data to View Number of the Lines ccccceeeeeeeceeeeeeeeeeeeeeeeeeeeeeeeeseeaeeeeeeeas 29 4 1 3 Details of the Correlation Data Displayed 00000nnnnnn000nnnnnennnnnnnnnnnnsrnnnnnnnnsennnnrrennsnnnrrreenene 30 4 1 4 Displaying densities of the Correlation Lines on the Bird s Eye Ma p ccccceeseeeeeeeeeeees 31 4 1 5 Uploading Users Own Correlation Data ccccccccceccccceeeeeeeseeeeeeeeeseeeeeeceeeeeesaeaeeeeeeeessaaaaeees 32 C KaPPA View4 Manual for Beginners 4 1 6 Format of the Correlation Data cccccccccccccccceeeeeeeeeeeceeeeeeaeeeeeeeeeeesaeeeeeeeeeeessaeaseeeeeesessnaaeess 33 ig ee 10 8 oe e S ee ee ee er eee E 34 4 3 Creation of Multiple Map ccccccccsesecceceeeeceeeeeeeeeeeeeseeseeeesseaceeseaeeessaaeeesseeeesseaaes 37 4 4 Utilization of User Maps ccccccssssecceecceeeeeeeceeessceecceeueeeecceeaageesessauseeeesseaaseeesesanaaaes 39 5 Oter FUNCIONS oo caresses ces on seuss ese annae EEEa aE ENE OOE ESEA 43 5 1 Universal Map MOJE corr EEE EE 43 5 2 Comparing two experiments in a SPECICS ccccccseeeeeeeeeeeeeeeeeeeeeeeeeeeeeaaaaseeeeeeeeeeee
40. elab ico Diver d Ad the daph data you aed to ee ard the inforrabon thes that the KaP PA enw peT is mfemeg con be dowrtoaded tom ere Coreg O AAAA Furuse DMA Research becitube Al Paghis Aiesered On the top of the window the main menu is placed a KaPPA View4 Manual for Beginners Main Temporary Upload Analysis Map View search Download e Main Returns to the main window Temporary Upload Uploading your experimental data User Map data and correlation data for your analyses is operated through this menu All the uploaded data is going to be deleted completely after you are logging out Analysis Uploaded data is displayed on the pathway maps through this menu It serves the central function of KaPPA View4 Map View You can browse plain pathway maps with no data from here Search You can search genes metabolites and enzyme reactions from here and access to the pathway maps which they are on Homology search function by blast to find genes 1s also provided Download The default experimental data publicly available on the KaPPA View4 and information data for genes metabolites reactions and maps for each species are downloadable as text files 2 3 Logoff You can logoff from the system by clicking Log off on top right of the main window Online Help amp Create Account e Log off y If you don t do any operations for 60 min after you log in the system regards as you are log off You are
41. ferase ae Vali lt i een Leucine Valine and ae L Isoleucine isoleucine metabolism ee 2 Uni00009 2008 03 28 L Leucine The experiment values are represented in this figure as colors of the symbols The correlation lines can be represented too when users are browsing the plain maps without experimental data 4 1 2 Filtering the Data to view Range of the Correlation Values The correlation data currently selected was calculated from 1388 Arabidopsis GeneChips Affymetrix and the gene to gene relationships which showed more than or equal to 0 6 of Pearson s Correlation Coefficients were included When the users would like to focus on much stronger relationships they can filter the data by the correlation values Please enter 0 9 in the left hand field of the Range column Click the Update Correlation button 28 KaPPA View4 Manual for Beginners Color Range Number Then only the lines having values between 0 9 and 1 0 are displayed Leucine valine isoleucine and alanine biosynthesis d F Lysine threonine anc f N Glycolysis methionine biosynthesis Ketol acid reductoisomerase Acatolactate synthase Threonine dehydratase J 2 Oxobutancate 2 3 Dihydroxy isovalerate L Threonine 2 Acatolactate Dihydroxyacd Acato 5 5 dehydratase cetoladate synthase Branched chain amino acid aminotransferase Alani aminotransferase 2 Aceto
42. h Player plug in ver 9 or higher is required to display the pathway maps it 1s already installed in your browser in the most cases If your browser doesn t have it please install it according to the following site Cu KaPPA View4 Manual for Beginners http www adobe com products flashplayer The operation of KaPPA View4 was tested in the following settings Windows XP Vista 7 Microsoft Mac OS X Apple Browser Internet Explorer 6 7 8 Safari 4 0 4 Mozilla Firefox 3 0 10 3 5 2 3 6 10 Mozilla Firefox 3 5 6 Google Chrome 3 0 Opera 9 63 10 10 In the case of Opera on Mac OS X the full screen view of the maps does not work Disable the pop up blocking function of your browser It is enabled for Safari and Opera in default 1 4 Other Manuals A full guide of all operations of KaPPA View4 is written in Manual for Users The procedure to create User Maps using free software Inkscape is described in detail in Manual of User Map Creation These manuals are also available form the top page of the KaPPA View4 site The Manual for Users and the Manual of User Map Creation are under preparation They will be available in 2010 2 Starting the Analysis 2 Starting the Analysis Login and Upload of Experimental Data For the first step of the analyses we introduce the way to login main menu and how to upload the experimental data to KaPPA View4 2 1 Login Visit to the following URL and
43. l is displayed By clicking this the symbols of the genes are shown in a pop up window 3 2 2 Colors of the symbols The symbol colors correspond to the values of the elements Click on the Color Legend at the bottom of the window Element List Correlation List Histograny Color ep Download Plain Map Print Map Copyright 2004 2009 Kazusa DNA Research Institute All Rights Reserved Relationships between the colors and the ranges of the values of transcript and metabolite changes are displayed in a small window Pe hoePP AS Veet Color Legend Wind ma rj hip karups or io egvdicole Lacersd index actin exc O KaFPA Virmi Color Lrernd Wind E x E Mipi kanman pipri ones Legend index action Color Legend Color Legand Trangezipa Metabolnal mscipl Melabolle log Ratio Ratio Linear Scale Lower Upper Color Lower Upper Colar 0 699 000 1 655 5 45 O l ji O07 25 21 1x iy I AN 3 Data Analysis Basic In this example the gene symbols are drawn in red when the log10 ratio of them in the comparison of the experiments was greater than or equal to 0 699 5 fold change As shown here the up regulated genes and the increased metabolites are drawn in reddish color the down regulated genes and decreased metabolites are in greenish color and the genes and metabolites of no change are in yellow The limit value for the strongest c
44. lied that the pathway was activated because expressions of a large proportion of the genes in the pathway were up When more than one Compared Experiment Pair have been set in the analysis a pull down list is displayed at the bottom left of the Bird s Eye map You can select the data to view here Eze KaPPA View4 Manual for Beginners Compared Experiment Ath 1 Select Displaying Correlation Data In the case Correlation is selected for the display mode densities of the correlations on the maps are displayed in the bars Details are described later 4 1 Displaying the Correlation Data ns Ta 4 Data Analysis Advanced 4 Data Analysis Advanced Various functions for analyzing omics data based on the pathway maps are implemented in KaPPA View4 Each of them is described in this chapter Combinations of the functions will provide new points of view for decoding omics data 4 1 Displaying the Correlation Data In the recent years a huge number of microarray data are available on public and it contributes to generate co expression data as a novel data resource A group of genes which are involved in a certain biological system could be expressed in coordinate manner throughout various conditions Therefore focusing on the unknown genes which co expressing with well known genes could give a hint to uncover the functions of the unknown genes ATTED II http atted jp for example is one of vanguards
45. lycarate A aalas 2 poh Polysaccharide metat CLA Miscellaneous carboh Amino acid nucleic acid Of Glyceraldehyde 3 phcephate cam dehydrogenase NADP phoephorylating Ribulcee bisphosphate p carboxylase small subunit Dihydroxyacetone gt Create Simple Map bisphosphate aldolase Aldolase Carton dixide Carbon dioxide Transketolase D Ribulose Atmosphere 1 5 bisphosphate D Fructoge 6 phoephate D Xyluloge Phosphcribulckinase 5 phoephate D Fructose 1 6 bisphosphate D Ribuloge 5 000 phosphate Fructoee bisphosphatase D Ribuloge 5 phosphate S epimerase D Erythroee 4 phosphate As shown here you can browse the transcriptome and metabolome data by selecting the species and the pathway maps 3 2 1 Symbols on the pathway maps The elements such as genes and metabolites on the pathway maps are represented as symbols below genes O Squares metabolites enzyme O Grete The color of the arrows correspond to KaPPA View4 Manual for Beginners reactions the mean value of the genes assigned to the reactions links to the By clicking the corresponding pathway name other maps pathway maps is displayed round rectangles with a text When there is not an enough space to Poe draw all the genes near by the Squares with enzyme reactions this symbo
46. m Arabidopsis So select an Arabidopsis thaliana AGI codes from the Array Type pull down list Select OOo Arabidopsis thaliana otus japonicus Agilent Kazusa Oryza sativa Agilent G4138A Oryza sativa AK Solanum lycopersicum Affymetrix AGI codes Press the Submit button to start uploading and registration the data to KaPPA View4 server After completing the process you can see the following message It takes a few tens of seconds The operation succeeded 2 4 2 Uploading of Metabolite Data Here we demonstrate the way to upload a metabolite data file It is similar to the case of DNA microarry data 2 4 1 Click the Temporary Upload from the main menu and select a file named Sample Ath _met_v csv in the data_metabolome folder in the sample data Temporary Upload Experiment User Map Correlation Experiment File Ae Upload The following window will open after clicking the Upload button aperenient ot Map Eim Lalit Selecl Expenment Mame Reprtiion Henber Cormont campie Aah A Sample Aih Bi i 2 Sample Ath C i Sample Al 1 Control Submit Preview a o 2 Starting the Analysis As this sample data is obtained from Arabidopsis select Arabidopsis thaliana from the Species pull down list In actual the sample data was generated by computational calculation and it did not contain experimental data of the real worl
47. mics data on the pathway maps with KaPPA View4 users have to login to the system and upload data files through the KaPPA View4 web user interfaces The POST data transferring function POST function provides logging in and data uploading environments through computational procedures without user s manual operation Therefore the developers can place for example View buttons in their database sites to view the data directly on KaPPA View4 ns 1 2 5 Other Functions You can find an example of the POST function in the post folder of the sample data In this sample transfer of a formatted data file is performed by post method of HTML FORM Open the file ExamplePostForm_v html with your Internet Browser p r ra TES F e CWDocuments and Settings tach we A C Documents and Settras eakura y AObo7 Ysa An example of POST data transfering function Select a file saved in the format for the KaPPA View POST data If it is accepted an URL will be displayed on the browser Accersing to the URL you can see the results on KaPPA Viewsd ore KaPPA View4 Y V4 We 8 100K Select a sample file Sample_postData txt by the Browse button and then click the Submit button If KaPPA View4 successfully accept the request an URL will be returned In the HTML sample the URL is displayed on the browser window fe htlpy hp kautanga hoe upload acin gt Wrdoerns Pilennel ie G je hrpe kp kausa ne jp kp
48. n check the format of the file Ate0i060 Atze46930 0 855 tle0io60 At3g09600 0 813 AtelM060 Atde 38960 0 8 tle0iog0 At3g48730 0 9 AteQi080 At5e55220 0 893 6 Atle 1080 At3g29185 0 891 7 Atle 1080 At4e29060 0 83 8 AtleQi080 Atles2990 0 889 9 jatleQ1080 AtleD5190 0 886 10 AtleQioso Atle f9B50 0 582 tle0iog0 At2g37660 0 881 AteQi080 Atles4510 0 88 AteQiO80 At5e13510 0 88 Ate0i080 Athed7190 0 877 1R ote AAN Aiar RR AIN In the case of gene correlation data a row consists of three columns 1 e gene ID gene ID and value correlation coefficient In the case of metabolite correlation a row contains compound ID compound ID and value correlation coefficient The data file must be saved in a comma separated vector CSV format 4 2 Simple Map As KaPPA View represents the gene expression data in the symbols on the pathway maps the genes not drawn on the maps cannot be analyzed While the microarray data and co expression data include a lot of non metabolic genes and unknown genes To expand the target genes for the analysis KaPPA View4 serves a function creating a Simple Map according to the gene ID list users input Square symbols for the genes in the ID list are arrayed on the simple map Myb Family AGRIS BRE BRB RBEBEBBBRORBBOOREBEBBRBBBORBDRBHBRORBRBERREE EE ES ATE S E ETR E ERB EBBRBHBBOROREBREBE E OBE BBR HRRBBHBROBDOOAB BHOBBROBEBRBEBEBHERRORBBBERBOOBRBRRBRSBSHREBEBBOABAOB
49. nes can be displayed on the maps This is the distinctive feature of KaPPA View and will help you for example to analyze the relationships between metabolic genes and transcription factors that control their expressions miasma Be ai The GS GOGAT Cute m 4 XY a a Ss ja a es L a at in Z n ree a Gina KaPPA View4 can handle multiple species and genes of several species can be displayed side by side on the maps The system also provides functions to upload and view the omics data from external applications ns Te 1 Introduction 1 2 Downloading Sample Data This manual guides practical operations of the system using sample data sampleFiles zip which is available from the top page of KaPPA View4 http kpv kazusa or jp kpv4 or form the main window displayed just after logging in KaPPA View 4 As the file is compressed as ZIP decompress the file with proper application before use data_metabolome Odata transcriptome post GzimpleMap URL link user Map E Readme EM tot E Feadme_dF txt The resulted folder contains several sub folders Please refer to the Readme_EN txt for their brief introductions 1 3 User Setup As KaPPA View4 is a web based system it works well with major web browsers Microsoft InternetExplorer Firefox Google Chrome Safari and Opera on any operating systems Windows XP Vista 7 Mac OS X and Linux Although the Adobe Flas
50. olor could be changed in the Histogram Element List Correlation List olor Legend Download Plain Map Print Map Real EE ee E bor hes Em iz fm rw if N oradan anant ea as e ee ad PARA h ap i i um Input the limit value in the Highest Linear Value field By clicking the Calc button you can check the frequency distribution of the elements in each color Push the Submit button to fix the setting Please refer to the Manual for Users for details 3 2 3 Switching the Compared Experiment Pair In the operation so far we selected two Compared Experiment Pairs namely Ath 1 and Ath 2 The data which you are currently browsing is checked in the upper control panel In the figure below the data from Ath 1 is represented KaPPA View4 Manual for Beginners Ath 1 Show All Experiments Transcript Sample Ath A Sample Ath B Metabolite Sample Ath A Sample Ath B Calvin cycle Phosphoglycerate kinase 3 Phoepho D glycerate Gluconeogenesis P ry Glycolate pathway 3 Phoepho D glycaroyl ome rn Triceaphosphate isomerase Glyceraldehyde prosp m 3 phosphate m m Dihydroxyacetone Glyceraldehyde 3 phoephate pheephate dehydrogenase NADP phoephorylating H Ribulcee bisphosphate carboxylase small subunit BBB rructee hon toda Transketolase Atmosphere bisphosphate aldolase Carbon di ide D Ribulose 1 5 bisphosphate
51. om the Display Mode Display Mode Name z Displaying the Map Names When the Name is selected for Display Mode the names of the pathway maps are displayed in the bars CO2 fixation and central carbohydrate metabolism calvin cycle _Glycolate pathway Glycolysisigluconeogenesis d Phoephoenolpyruvate and pyruvate metabolism TCA oyda i hlarsedebe cca Displaying the Experiment Data a Tz 3 Data Analysis Basic When you select Experiment the bars turn to as below 102 fixation and cantra T 251 25 ts 7172 M T 3535 D N M1595 T5131 B T i 1607 T4646 ET t i EE 7 73 73 M a a m aa carbohydrate metabolism T 412 D 4 j OO E o s U U T 100 B e E The color gradation represents followings Ex T azas M e ooo T Transcripts The denominator shows the number of the genes drawn on the pathway map 43 genes The numerator indicates the number of the genes having valid values in the current data 42 genes values for one gene was invalid M Metabolites The valid metabolite number numerator 8 and the metabolites drawn on the map denominator 14 The numerator and denominator values at the pathway categories are the compiled values of all the pathways included in the category Color gradation of the bar The proportion of the genes or metabolites painted in the color Therefore if the bar for transcript was strongly painted in red it imp
52. omerase Fructose bisohosohatase aldolase When Universal is selected the button is not displayed Please specify a species A following window will open Simple Map Create New Simple Map Created Maps Map Name Gene List Add Clear Load From File Load From Map Redraw Let s create a Simple Map from gene ID list in the sample data Push the Browse button and select a file named myb_agris txt in the simpleMap folder By clicking the Load From File button the gene IDs written in the file is read and displayed in the Gene List area 35 4 Data Analysis Advanced New Simple Map Map Name Gene List At3q27920 At3q09230 At3g12820 At2g25230 At2q32460 Atic69560 At3q01140 cear You can input the gene IDs directly in the Gene List area By clicking the Load From Map the gene IDs on the pathway map currently viewed is displayed in the Gene List area Input Myb Family AGRIS as Map Name and press Add button New Simple Map Map Name Myb Family AGRIS Gene List Then the registered map name is appeared in the Created Maps area Created Maps Myb Family AGRIS Click the Redraw button to redraw the pathway tree in the main window Finally close the window The registered map is appeared under the new branch named Simple Map in the pathway tree You can use it for your analyses same as the default maps
53. press Guest Login http kpv kazusa or jp kpv4 Welcome to KaPPA Views Classic Go to My Page Nae Enter Password Please cick the nate tana Aas date is treaty mares in we Dy vrs Uor e a0vENCed pow Can Save your own data sore aang pour acc REPY EREE ieri pe yeere ate Ost ahon ree Gene wtormation for Oryz anged August 15 2010 The transe areni ade veir ybg boaten sed tor gene mates a We changed the gene DOs oft to the lotus krs from RAP DB s that the cormelation data of nce A ba t represented The old settings are preserved as the species name Oryza satha RAP OG transcegt so your mecroatay Gata descnbed as the AEDE lOs can be analyzed wth A The dats also can be analyzed in the updated Oryza sata by selecting the array type TRAP DB transcnpty 2 2 The Main Menu After logging in you can see the main window Tempi Uplowd Tou can upload your can data jexpecmeet data map date and conelation tia tor Mayes Other users DEAR ATAR ba JOERS DI Them Alber heer adn here KP PA Gite ocr th tow athe eed ta 3 t be chee bom Pie tarai Qn che daah of be dia brna pihassa book a thee tangia Glew Anahpen Vou Can Create baril expenirend beta here ti view the dla on the eplabobe palha ipri Mig Wier A he melatobe paltry mapi can Be Groene Fon hra Trach Geren nebaboiiys aed erry irabaled in Kal A Ven are nearchable anc you can check whai metab mapa they are diwan on o soma SPRIE bigest search EHE EMH earch i
54. sketolase Select Species Element List Download Plz A pop up window will be displayed Check on the species you would like to display on the maps and press the Submit button and then Redraw button to refresh the map on the main window l Kaf TA Wiel SpPeties Select Wndoss Internat Lpkeer 1 fe hey Kaaba ee ole po lipat aeaii a 1 a mt rl Eli E i RIARTE HN E CETTE T Eer TOLI E era KaPPA View4 Manual for Beginners In the Universal Map Mode you can compare omics data between the species too When you set more than two Compared Experiment Sets originated from several species an extra button Show All will be displayed on the top right of the pathway map Click the button kiniad Eparnents Ath Datal Sly Data Ath Data Ath Data Show All Tranecripa sampa _AN Leaves 21 days sample_As TS cultured cells 14 days Metabolite Kazusa TS GewihCune day 017 Kkazusa TAT GrewthCune day 02 Calvin cycle p n a a aan Gluconespenesis bra Ma e d Phea 2 Phaea D giomi TEE A TRBepTephane SETHA z Drane mzs gt phei E saa epee Giyeninin pathwny ah Dryga h J Gimdo phowsten oneaphuna deol rpcbeger G d pe NAD Wehnaphoniatng pu Ribuioss biaphanphana OTOA urd mun You can select one data from each species and click Submit button Compared Experiments Ath Datal Sly Data Ath Data Arabidopsis thaliana Solanum lycopersicum
55. sseaes 10 3 Data Analysis BASIC sicssncsiveciacencadeninsvenunddanertenarensstenienesnssexnrediecerewaneesvieessanennes 13 3 1 Data selection for browsing ccccccesseeecceeesceceeseeecsaeeecsauseeeseseeeesaeeeessauseessenseesseaes 13 3 2 Data browsing on the MAPS ccccccsescecceeeseeceeeseecceuseeecsauseeceauseeeeseeeessagseessanseesseass 17 3 2 1 Symbols on the pathway maps ccccccsssseccccceesececceeeeeeecceeeseeecseeaeeeeseeageeeesseageeessseseeessnas 18 3 2 2 Colors df ihe SVIMDOIS escrire EE E E 19 3 2 3 Switching the Compared Experiment Pai cccccccccccccceccseseeeeeeeeeeeeeeeeeeeesesaeeeeeeeeeeessaaeeess 20 3 2 4 Details of genes metabolites and enzyme reactions ceccccseeeeeceeeeeeeeeeeeeeeeesaeeeeeseeeeens 21 3 3 The Bird s Eye Map ccccccccsesccccessseeeseeeeeeseaseeeeeeseeessaaeeeeesaeeeeseaseeessaeeeessaseesasaeeesaass 22 Displaying the Map Names ccccccsscecceeseecceeseecseueeeceaeeeeseseeeeeeeeseasesseseessaueeecsuseesseueeessageesssags 23 Displaying the Experiment Data cccccseececceeeeeceeeeeeceeeeeeseeeeeeeeeeeeeeeeeessaeeeesaueeessaueeeseeeeeeaeeeesaess 23 Displaying Correlation Data vcsscncsseaceccteseswonsicenecvaschassmaceeetesuennennesteesadiensaness nactubasierascoeeteeeaceacseetuenaee 25 4 Data An lysis Advanced seesinane tuaana anaa teases 26 4 1 Displaying the Correlation Data ccccccccccccccsceeeesssseeeeeeeseaeeessee
56. yd upland act baie i ea E i A w g 2 bps kp kannar ping up ed action Fama TT ea om y Cut and paste the URL in the address field and jump to the site lt Je v JE http kpv kazusa or jp kpv4 redirectaction lid ab14740e 4b15 434a 8b02 4d8fb16b6f9d referId ze4a3985b 25ad 48b7 9fcc caa9e1f4d e The result will be displayed on the browser window aas KaPPA View4 Manual for Beginners Example of POST transfered Map Giycerol metabolism Kikoin Caio Semple Map Oo A nd Rated Map Upeaie Conii User Map data could be included in the POST data too After jumping to the KaPPA View4 users can continue to browse the data and start next analysis with the posted data Details on the transferring procedure sample code of PHP and the format of the data file are described in the Manual for Users 6 User Account 6 User Accounts After logging in to the KaPPA View4 system as a common user Guest User you can create freely your own account with a simple procedure Power User If you get your account you can save your own experiment data correlation data and User Map data on KaPPA View4 It helps you to start the analyses immediately after logging in There are no differences on the analysis functions between the user authorities 6 1 Creating an Account Log in as guest KaPPA View 4 Kazusa Plant Pathway Viewer Login Enter with your name and password Enter as a guest Name
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