Blixem User Manual
Contents
1. By average similarity by Blosum62 By percent identity By percent identity Blosum62 By residue Erik s By residue Toby s E Finished calculating tree i Edit current colour scheme Save colour scheme Load colour scheme Figure 12 Menu tear offs File menu Wrap for printing Print Show tree 36 Tree settings Recalculate tree Show conservation plot la Save Ctrl s leq Save as Shift Ctrl 5 Output score coords Fetch sequences via WWW Compare all and output identities amp Clean up windows Figure 13 File menu Quit Quit Belvu close all windows and exit Wrap for printing Open a window showing a wrapped alignment suitable for printing Print Print the current window note that you should use the print 16 Show tree Tree settings Recalculate tree Show conservation plot Save Save as Output score coords Fetch sequences via WWW Compare all and output identities Clean up windows Edit menu option from the wrapped alignment window to print the wrapped view Open the tree window calculates the tree if it has not yet been calculated Edit the settings used to calculate and display the tree Use this to recalculate the tree after making changes that invalidate it e g deleting rows Show the conservation plot window Save the alignment in the current format Save the alignment allows you to select a different file format and choose whether coordinates shou2. Show all lines that were previously hidden By residue Erik s By residue Toby s By residue Cys Gly Pro By residue Cys Gly Pro His By residue Clean slate By residue Custom By average similarity by Blosum62 By percent identity By percent identity Blosumb2 FL Edit current colour scheme Save colour scheme Load colour scheme Figure 15 Color menu Erik s Toby s Cys Gly Pro Cys Gly Pro His Clean slate Use Erik s original built in residue colour scheme Another built in residue colour scheme A colour by residue scheme where only cystine glycine and proline are highlighted A colour by residue scheme where only cystine glycine proline and histidine are highlighted Clear all colours used for when you want to create a new colour scheme starting with all colours being white 19 Custom By average similarity by Blosum62 By percent identity By percent identity Blosum62 Edit current colour scheme Save colour scheme Load colour scheme Settings menu This option will become enabled when a residue colour scheme has been customised by editing it or loading it from file if you change to a different colour scheme you can toggle back to the custom colour scheme by selecting this option A colour by conservation scheme colouring by average similarity by Blosum62 A colour by conservation scheme colouring by percent identity A colour by conservation scheme colouring by both pe
3. branch lengths Display organism Interactions Action when picking a node Swap Es S Cancel af Apply Figure 8 Tree settings dialog The options are as follows Note that changing the tree building method or distance correction method will force the tree to be recalculated which may take a long time for large alignments Tree building method Choose whether the tree should be built using the neighbour joining or UPGMA method Distance correction Select the distance correction method to use method Tree scale Adjust the horizontal scale used to draw the tree set a smaller number to decrease the width of the tree or a larger number to Increase It Line width Set the line width to use for the branches 0 1 gt 1 pixel Display branch lengths Whether to label branches with their lengths Display organism Whether to display the organism next to the sequence name Action when picking a Swap when you click a branch its two child nodes will be node swapped 13 Reroot when you click a branch the tree will be re rooted with that node as the root Note to revert to the original tree select the Recalculate tree option from the right click menu Conservation plot To display the conservation profile select Show conservation plot from the File menu The conservation profile window will open displaying a plot of the conservation vertical axis against the column numbers horizontal axis The average conservation i
4. the File menu Note that menus with a dotted line at the top can be torn off by clicking on the dotted line A torn off menu will stay visible on top of the Belvu window and can be repositioned by dragging its header bar Click the dotted line again to get rid of it 15 a0 File Edit EHA Settings Sort Help H G T A AA Colum 57 Q21691_CAEEL 673 1001 G 718 1 match 21x412 40 60 80 90 By residue Erik s YQ53_CAEE Fig ce S IGLOTIQLCQQTVDKM GQGGRQTID FFVEIPNAVRGKAVC Q21691 CAE beberle ten y s LGQQTIQISSETADKF DHGGKQTID P FLIDVPESVGHRVVC 048771_ARA By residue Cys Gly Pro LLGIEYSYNIPLI TAG76_CAE is SRIVADKITNKYL 016720 CAR co ILLPNVRPR 1 PINH_ARAT By residue Custom IGGRNT VLVDAISCRIPLV AGO1_SCHPQ E SEPARE 076922 prq By average similarity by Blosum62 PIWI_DROMW By percent identity Q17567_CAB By percent identity Blosum62 PIWL1_HUMA PIWI_ARCFU M Edit current colour scheme Y1321 MET Save colour scheme 067434_AQ 921495 cag Load colour scheme 016386_CAEEL 548 847 QLIMFITK SMNN WATEL By residue Cys Gly Pro 002095_CAEEL 574 878 QLLFFVVK SRYN YHQQIKA By residue Cys Gly Pro His Q19645_CAEEL 674 996 PFWLFISD DVPN IHECLKFEERMSDIPTOHVLLKNVKKMRDN IEKK SQGGRRAYDLTLDN IVMKAN I By residue Clean slate Q23415_CAEEL 40 350 KFAFVITD DSITH LHKKYKALEQKSMMVI VK DGKRLTLENVINKTN US Gall E A 062275_CAEEL 594 924 TFWFIITD DSITT LHQRYKMIEKDTKMI INAGKR LTLENVIN Y
5. the sequences in unaligned Fasta format i e with gaps removed Raw The raw file format is as follows Raw files must be passed using the command line argument because Belvu cannot detect this file format automatically lt name gt lt sequence gt lt name gt lt sequence gt The Belvu Windows Main window The main Belvu window contains the alignments Residues are coloured by conservation or by residue type use the Color menu to change the colour scheme 21x412 10 60 80 90 YQ53_CAEEL 650 DILVGIAR EKKPD VHDIL MG GQGGRQTIDNVMR FFVEIPNAVRGKAVC Q21691_CAEEL 673 TIVFGIIA EKRPD MHDIL MR DHGGKQTIDNVIR FLIDVPESVGHRVVC 048771_ARATH 542 FILCILPERKTSDI YGPWK Q9ZVD5_ARATH 577 FILCVLPDKKNSDL S MLSVERTPAFTVI TAG76_CAEEL 660 CIIVVLQS KNSDI SRIVADKITNKYL 016720 _CAEEL 566 LIVVVLPG KTPI ILLPNVRPR 1 PINH_ARATH 625 LLLAILPD NNGSL VLVDAISCRIPLV AGO1_ SCHPO 500 YLFFILDK NSPEP SLIPKSNP L 076922_DROME 555 IVMVVMRS PNEEK PIWI_DROME 538 LILCLVPN DNAER Q17567_CAEEL 397 MLVVMLAD DNKTR PIWL1 HUMAN 555 IVVCLLSS NRKDK cas PIWI_ARCFU 110 GIMLVLPE YNTPL PWILNVDPEK Y1321_ METJA 426 CFALIIGKEKYKDND FILDSKTPYDY 067434 AQUAE 419 LVIVFLEEYPKVDP YKLKETEGKVDA Q21495_CAEEL 52 GIVLPTPRIFFRDG 016386 _CAEEL 548 QLIMFITK SMNN 002095 CAEEL 574 QLLFFVVK SRYN YHQQI i Q19645_CAEEL 674 PFVLFISD DVPN IHECLKFEERMSDIP YT ADIPRDLACW Q23415_CAEEL 40 KFAFVITD DSITH LHKK L
6. Belvu User Manual Written by Gemma Barson lt gb 10 sanger ac uk gt Wellcome Trust Sanger Institute 22 September 2011 Revision History Revision Date Author First revision Belvu v4 4 1 18 01 11 Gemma Barson Updated for version 4 14 02 07 12 Gemma Barson Contents REVISION HISTON Y simi Gio e a ees eee GOtUna ee Rea Pan Hes VW steep ccc statsascricacteretcaia ales E A E et da eased oe A 5 NS MNS ec teers os essere sts echo oe eects octet de ease A 5 The Belvu WindOWS ccccccccocoocooooccoo ooo S ME RU E E AA 7 SOLOING A E E E E EE EE A 8 PE SCC E A ida 9 TOO OBE AA AI IU UDI A 9 A E E wie ag a te ec ec tetas ea dees a edo dA veo NEEE alsa rv cated 10 Mi aoc ENAN EE cc ee nda E E sete TE sw cei seuss vets Sn eae Gace atone one dd DB TIS VINO Mide L DTS r A PETEA AI o I II un La Conservation placita 14 Conserva On DO ICI aro ANOS ar 14 OMS CEVA OM DIOS IM air iia 15 MA Mortis EME Malasia ocre EEEE EEE 16 OS A 17 Ps A oeddwensansanaceewetiesde end cetaceusesandeetwetmaseoataeswennseuwndeeeosnt 19 Seas E o A T E E 20 HO Mini aii 21 Keyboard SNOFIGUtS iia redone Introduction This manual explains how to configure run and use Belvu Belvu is a multiple sequence alignment viewer and phylogenetic tool It has an extensive set of user configurable modes to color residues by conservation or by residue type and some basic alignment editin
7. EQKSMMVI YTVSDAKKSMT 062275 _CAEEL 594 TFWFIITD DSITT LHORYKMIEKDTKMI YVFVDAKKQL Figure 1 Alignment window in colour by conservation mode ANO Belvu PF02171 seed sele et zi i E Q Q a Column 67 Q21691_CAEEL 673 1001 K 728 1 match E ui YQ53_CAEEL Q21691_CAEEL 048771_ARATH Q9ZVD5_ARATH TAG76_CAEEL 016720_CAEEL PINH_ARATH AGO1_SCHPO 076922 _DROME PIWI DROME Q17567_CAEEL PTWL1_ HUMAN PIWI_ARCFU 062275 CAEEL Figure 2 Alignment window in colour by residue mode At the top of the alignment list is a header displaying the number of sequences and alignment length e g 21412 means there are 21 sequences and the alignment length is 412 The alignment list contains the following columns Name The sequence name Start The start coordinate in the match sequence End The end coordinate in the match sequence Score Only displayed if a scores file was loaded displays the score of the sequence Sequence Displays the sequence data Selections Click on a row to select that alignment Details about the selected row will be shown in the feedback box on the toolbar If there are other sequences with the same name their names will be highlighted in the alignment list but only the clicked row will have the whole row highlighted The number of matches is shown in brackets in the feedback box If you clicked within the sequence area a column will also be selected the column number 1 based from
8. dotter lt alignment_file gt where lt alignment_file gt is a file or pipe containing the multiple alignment in Stockholm Selex MSF or aligned Fasta format see below Run belvu without any arguments to see brief usage information or for more detailed help run belvu help File formats Belvu currently supports Stockholm Mul Pfam Selex MSF and aligned and unaligned Fasta formats Belvu will automatically detect which file format is supplied The raw file format can also be used but you must pass a raw file using the r argument because Belvu cannot detect this format automatically Selex Selex is the native format used by Sean Eddy s HMM package HMMER For details see http www psc edu general software packages hmmer manual node46 html Each line contains a name followed by the aligned sequence A space dash underscore or period denotes a gap If the alignment is too long to fit on one line the alignment is split into multiple blocks separated by blank lines The number of sequences their order and their names must be the same in every block even if a sequence has no residues in a given block Other blank lines are ignored You can add comments to the file on lines starting with a seql ACGACGACGACG seq2 GGGAAAGG GA seg3 UUU AAAUUU A seql ACG seq2 AAGGG seg3 AA UUU Stockholm Also known as Mul or Pfam format Stockholm is the native format used by Pfam and Rfam to d
9. e was no previous search result Tree The tree window can be opened from the main window using the Show tree option on the File menu The tree window will show a distance based phylogenetic tree of the current alignment using the default settings To edit the tree settings 10 before calculating the tree first select the Tree settings option from the File menu Click on a sequence name to select a sequence in the tree the sequence will be highlighted in both the tree and the main window Click on a branch to either swap the nodes or re root the tree from that branch see the Tree settings section for more details ADO N Belvu Neighbor joining tree using Scoredist correction distances of PFO2171_seed selex YQ53 CAEEL 650 977 CAEEL Q21691 CAEEL 673 1001 CAEEL Q21495 CAFEL S2 336 CAEEL 016386 CAEEL 548 847 CAEEL 002095 CAEFEL S 4 878 CAEEL Q19645 CAFEL 674 996 CAEFEL 023415 CAEFEL 40 350 CAEEL 062275 CAFEL 594 924 CAEFEL 048771 ARATH 542 860 ARATH 09 7VD5 ARATH 5 7 885 ARATH PINH ARATH 625 946 ARATH AGO SCHPO S00 799 SCHPO TAG 76 CAEFEL 660 966 CAFEL 016 20 CAFEL 566 867 CAEFEL 076922 DROME 555 852 DROME PIWIL DROME 538 829 DROME Q1 567 CAEEL 397 708 CAEEL PIWL1_HUMAN 555 847 HUMAN PIWI_ARCFUNA1O 406 ARCFU Y1321_METJA 426 699 METJA 067434 AQUAE 419 694 AQUAE Figure 5 Tree window Tree menu The tree menu can be accessed by right clicking anywhere in the tree window Close The op
10. g capabilities It can generate distance matrices between sequences and construct distance based trees either graphically or as part of a phylogenetic software pipeline Key features include Residues can be coloured by conservation with user configurable cutoffs and colours Residues can be coloured by residue type user configurable Colour schemes can be imported or exported Swissprot or PIR entries can be fetched by double clicking The position in the alignment can be easily tracked Simple editing commands for rows and columns is supported although Belvu is not intended to be a full editor The alignment can be saved in Stockholm Selex MSF or FASTA format Distance matrices between sequences can be generated using a variety of distance metrics Distance matrices can be imported or exported Trees can be constructed based on various distance based tree reconstruction algorithms Trees can be saved in New Hampshire format Belvu can perform bootstrap phylogenetic reconstruction Belvu can be used as a graphical tree viewer or as a command line tool for use in phylogenetic software pipelines Belvu is maintained by the Wellcome Trust Sanger Institute and is available as part of the SeqTools package The software can be downloaded from the Sanger Institute s website http www sanger ac uk resources software seqtools Getting Started Running Belvu As a minimum Belvu takes the following required arguments
11. gure 17 Help menu Show the help pages Show the About dialog 21 Ctrl Ctrl Shift Ctrl Shift Ctrl PageUp PageDown Ctrl up Ctrl down Home End Ctrl W Ctrl Q Ctrl S Shift Ctrl S Ctrl P Ctrl H Ctrl F CtrI R Ctrl T t equal minus Old style Belvu shortcuts Left arrow Right arrow Ctrl left Ctrl right Up arrow Down arrow Insert Delete Keyboard shortcuts Recommended shortcuts consistent with other SeqTools programs Scroll one column left Scroll one column right Scroll one page left Scroll one page right Scroll to leftmost column Scroll to rightmost column Scroll one page up Scroll one page down Scroll one row up Scroll one row down Scroll to top of alignment list Scroll to bottom of alignment list Close the current window If this is the main window it quits the application Quit the application Save the alignment in the current format Save the alignment in a different format Print the current window Open the Help pages Find sequences Make non redundant Remove partial sequences Toggle between colour by residue and colour by conservation mode Zoom in Zoom out Scroll one page left Scroll one page right Scroll one column left Scroll one column right Scroll one page up Scroll one page down Scroll to leftmost column Scroll to rightmost column 22
12. isseminate protein and RNA sequence alignments The file must start with a line giving the format version and end with It has one domain per line STOCKHOLM 1 0 lt sequence_name gt lt start gt lt end gt lt sequence gt A The residues must be aligned and gaps should be represented by dots Markup lines can also be included see http en wikipedia org wiki Stockholm_ format for more details MSF Note on the MSF format The Check line has to come before the first line that does not start with a space The only legal exception is the line PileUp of from GCG programs Pileup lt filename gt MSF lt len gt Type lt type gt Check lt check gt Name lt namel gt Len lt len gt Check lt check gt Weight lt weight gt Name lt name2 gt Len lt len gt Check lt check gt Weight lt weight gt Fl lt namel gt lt sequence gt lt name2 gt lt sequence gt lt namel gt lt sequence gt lt name2 gt lt sequence gt The sequence names can include coordinates e g lt name gt lt start gt lt end gt Fasta In Fasta format the sequence name is on a line starting with gt gt and the sequence on the following line s Input files for Belvu must be in aligned Fasta format where gaps are included so that each sequence is the same length gt seql ACGACGACGACG ACG gt seq2 GGGAAAGG GA AAGGG Belvu does not accept unaligned Fasta files as input but can output
13. ld be saved and what separator character to use Only applicable if scores are loaded outputs the score and coordinates of the currently selected sequence to the terminal Enables fetching of sequences over HTTP Compares each sequence against each other and outputs their identity and score to the terminal along with some summary information about the maximum minimum and mean score and identity Close all windows opened by this instance of Belvu does not close the main window 17 Remove highlighted line Remove many sequences Remove gappy sequences Remove partial sequences Make non redundant Remove outliers We oe eo Ae ee A Y LL ol tad Remove columns 5 S lt Remove columns left of selection inclusive Remove columns right of selection inclusive gt Remove columns by conservation Remove gappy columns Automatically remove empty columns Read labels of highlighted sequence and spread them Select gap character Hide highlighted line Unhide all hidden lines Figure 14 Edit menu Remove highlighted line Remove many sequences Remove gappy sequences Remove partial sequences Make non redundant Remove outliers Remove sequences by score Remove columns Remove columns left of selection Remove the currently selected line Enables a mode where you can double click on sequences to remove them The cursor will change to indicate that you are in this mode Select the o
14. ption again press the Esc key or right click to cancel this mode Remove sequences that have more than a given percentage of gaps Removes partial sequences Remove sequences that are more than a given percentage identical to any other Remove sequences that are less than a given percentage identical to any other Only applicable if scores are loaded remove sequences that have a score lower than a given threshold Remove a specific range of columns Removes the columns to the left of the currently selected column which is displayed in the feedback box on the toolbar if a column is selected The operation is inclusive so the currently selected column will be removed as well 18 Remove columns right of selection Remove columns by conservation Remove gappy columns Automatically remove empty columns Read labels of highlighted sequence and spread them Select gap character Hide highlighted line Unhide all lines Color menu Removes the columns to the right of the currently selected column The operation is inclusive so the currently selected column will be removed as well Remove columns with a maximum conservation between specified values Remove columns with more than a given percentage of gaps After deleting sequences columns that are left empty are automatically removed if this option is enabled Undocumented Change the character used to display gaps in the alignment Hides the currently selected line
15. rcent identity and average similarity by Blosum62 Edit the current colour scheme If in colour by residue mode allows you to edit the residue colours if in colour by conservation mode allows you to edit the thresholds and colours for the different levels of conservation Only applicable in colour by residue mode save the current colour scheme to file Only applicable in colour by residue mode load a colour scheme from file Only colour residues above 1D tl ld Set 1D threshold Ignore gaps in conservation calculation Exclude highlighted from calculations Use gray shades for printing Display colors faster without Highlight lowercase characters Figure 16 Settings menu Only colour residues above ID threshold Ignore gaps in conservation calculation Exclude highlighted from calculations Use gray shades Only applicable in colour by residue mode only colour residues that have a percent identity above the threshold specified by the Set ID threshold menu option Only applicable in colour by conservation mode ignore gaps when calculating the conservation Exclude the currently selected row from colour calculations Only applicable to colour by conservation mode use grey shades suitable for printing 20 Display colours Whether to show colours or not faster without Highlight lowercase Highlights lowercase characters characters Help menu Help About T Help Ctrl H 7 About Fi
16. s shown as a red line Boo NI Belvu Conservation Profile Window 1 4 1 O i ll Fi i 40 5 El TI CL Figure 9 Conservation plot Conservation plot menu Right click anywhere on the conservation plot to display the menu a Close 2 Print to Plot settings Figure 10 Conservation plot menu The options are Close Close the conservation plot window Print Print the conservation plot Plot settings Show the plot settings dialog 14 Conservation plot settings Select the Plot settings option from the right click menu on the conservation plot to show the plot settings dialog Ao IN Belvu Plot Settings Window size fi l 10 Y scale 20 X scale Line width i X Cancel Figure 11 Conservation plot settings The options are Window size Specify the size of the sliding window used to smooth out the curve set a larger value for a smoother curve The minimum value is 1 which means no smoothing is done X scale Adjust the scale of the horizontal axis set a smaller value to compress the scale or a larger value to expand it Y scale Adjust the scale of the vertical axis set a smaller value to compress the scale or a larger value to expand it Line width Set the line width to use for the drawing in pixels Main menu The main menu can be accessed via the menu bar at the top of the main window Right clicking in the main window is a shortcut to
17. size in the alignment list Find Open the Find dialog The feedback area on the toolbar displays the following information Column lt column gt If a column is selected this displays the column number 1 based from the left most column lt name gt lt start gt lt end gt lf a sequence is selected this displays the sequence name and its start end coordinates lt residue gt lt coord gt If a column and sequence are selected this displays the residue and coordinate of that column within that sequence lt n gt match s If a sequence is selected this shows the number of sequences in the alignment with the same name 1 gt only the current sequence has that name Find dialog The Find dialog allows you to search for sequences by name Open it by clicking on the toolbar icon or by using the keyboard shortcut Ctrl F AOS Belvu Find sequences a Name search wildcards and 7 O _Geback cprormara cose Box Figure 4 Find sequences dialog Enter the text you wish to search for The text can include the wildcards for any amount of any character or for one occurrence of any character Hit OK to close the dialog and search If found the first matching result will be highlighted in the alignment list Alternatively click Forward or Back on the Find dialog to perform a search forwards or backwards from the last search result These operations will start from the beginning of the list if ther
18. the left that you clicked will be shown in the feedback box along with the residue and the sequence coordinate at that column for the selected sequence Middle click in the alignment in order to select a column the current column will be highlighted while the middle button remains pressed and you can drag to other columns to see column information dynamically When you release the mouse button the display will scroll so that it is centered on the selected column Fetching sequences Double click on a row in the alignment to fetch that sequence the program used to fetch sequences must be specified in the BELVU_FETCH environment variable before Belvu is opened e g in a C shell terminal setenv BELVU_FETCH pfetch F Toolbar The toolbar contains shortcuts to several of the menu items as well as a feedback area displaying information about the currently selected row and or column Hamal Q G_ column 57 Q21691_CAEEL 673 1001 G 718 1 match Figure 3 The toolbar The toolbar buttons are as follows Help Display the help pages See the Help menu Remove many sequences Start the mode that allows you to double click to remove sequences Click again or press Esc to cancel this mode See the Edit menu Edit current colour Edit the current colour scheme see the Color menu scheme Sort alphabetically Sort sequences by name see the Sort menu Zoom in Increase the font size in the alignment list Zoom out Decrease the font
19. tions on the tre print la Save Tree Tree settings Recalculate tree Find putative orthologs Show organisms Figure 6 Tree menu 11 Close Print Save Tree Tree settings Recalculate tree Find putative orthologs Show organisms Organisms window Close the tree window the tree will not be deleted and can be opened again without recalculating Print the tree window Save the tree in New Hampshire format Open the tree settings dialog Forces the tree to be recalculated this is required after the alignment has changed and the tree is now invalid e g if rows have been deleted Highlights putative orthologs in the tree and outputs their details to the terminal Opens a window showing the list of organisms and outputs the number of organisms to the terminal Select Show organisms from the right click menu in the tree to display the organisms window which lists all of the organisms in the alignment A X Belvu Organisms AQUAE ARATH ARCFU CAEEL DROME HUMAN METJA SCHPO Figure 7 Organisms window Tree settings To open the tree settings dialog use the Tree settings option from the File menu on the main window or from the right click menu on the tree window 12 ADO IN Belvu Tree Settings Build methods Tree building method NJ Display options Tree scale 0 30 Line width 0 30 L Display
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