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<text>Aligner is a freeware HyperCard 2.x stack for manual alignment of sequences by D. J. Eernisse, author of DNA Translator stack. Aligner was inspired by a stack called MultiDNA by Ralph Gonzalez, but was created completely from scratch. Enhancements compared to MultiDNA include: 1) Number of sequences that can be aligned increased from 6 to 100; 2) interleave formatting and number of lines automatically adjust for the number of sequences imported; 3) now accomodates sequences up to 30,000 bp or amino acid residues long; 4) imports or exports multiple sequences that are simple return-delineated strings or in 'DNA Translator' stack string format (i.e., 'Name<space(s)>AGCTGA...<rtn>'); 5) optionally interleaves exported sequences; 6) enlarged for use on a 640 x 480 standard color or larger monitor (automatically adjusts for standard small Mac screens); 7) much faster switching between sequences to be edited; 8) can toggle between match characters (dashes) to the first sequence and no match characters; 9) color-codes base pairs, amino acids, or codons, including support of alternative genetic codes; 10) speaks nucleotide/peptide characters during keyboard entry or after entry starting from the current insertion point; 11) various sequence manipulations, such as lower <-> UPPER case; 12) addition of gaps to all but the current sequence supported; 13) align amino acids, then introduce gaps to the corresponding unaligned nucleotide strings to preserve the amino acid alignment; 14) custom menus; 15) optional help facility displays the function of fields or buttons as the cursor enters them.The best way to get the most current version of Aligner or the more extensive DNA Translator stack package, is by anonymous ftp to "um.cc.umich.edu" then "cd gdef" and "get aligner.hqx" or "get dnastack.hqx" for Aligner stack only or the entire package, respectively. Then debinhex the resulting files. Aligner by itself is less than 300K uncompressed, while DNA Translator and associated files occupy about 1 MB on your hard disk or 500K in compressed Binhex format. This stack is only free for any noncommercial use and is not public domain. It is copyrighted 1991, 1992 by D. J. Eernisse, and uses some XFCN resources copyrighted by Nigel Perry with similar copyright restrictions (see stack script for details). For more information, contact: Douglas J. Eernisse, Museum of Zoology, Univ. of Michigan, Ann Arbor, MI 48109 USA, (usergdef@um.cc.umich.edu or usergdef@umichum.bitnet). ••••••••••••Scroll down for more information:Please cite as:Eernisse, D.J. 1992. DNA Translator and Aligner: HyperCard utilities to aid phylogeneticanalysis of molecules. CABIOS 8(2): 177-184.or:Eernisse, D.J. 1991. Aligner: HyperCard stack for manual sequence alignment. Electronically published software available by anonymous ftp to Ftp.Bio.Indiana.Edu.To display or hide a baloon-style help facility, click on the question mark icon. You probably will find it better to turn off this help facility if you are doing much editing.To get started, use the provided sample data, your favorite word processor, or DNA Translator stack to type in some simple "string" sequences of the form:name1 ATGTACGT…name2 ATGGACGT…name3 ATGTTACGT…name4 ATGTTACGT…The name can vary in length but cannot have internal spaces and should be followed by one or more spaces to separate it from the sequence, which also cannot have any internal spaces. The name and entire sequence should be on a single line. Valid nucleotide or amino acid symbols are supported in either upper or lower case. If you want to import sequences that are in interleaved or a number of other standard formats, use DNA Translator to convert them to "string" format first. The use of a name preceding each sequence provides convenient compatibility with DNA Translator but is not required. You can skip names if you wish and simply have one long sequence per line. The file to be imported should be saved in text (ASCII) format. Aligner sometimes will automatically add default names (e.g., seq1, seq2, ...) if it needs them for special operations. You can have up to 100 sequences, each up to 30,000 characters, although stack operations slow down well before these limits are approached.Once you have a disk file with "string" sequences, use "Import Multiple" from the "File" menu, which will prompt you with a standard "Open File" dialog box. The rest of importing is automatic. Export sequences in a similar manner.If you want to import sequences in FASTA or Intelligenetics format, the most recent version of DNA Translator will convert from these formats to the "Translator" named sequence string format required. Likewise, you can convert back to these formats if you want. If there is ever large demand, I will include these conversions within Aligner. GCG users can use "TOIG" or "FROMIG" to convert to Intelligenetics format. You will need to become familiar with the scroll window and the up/down interleave scroller. An option-click on the up or down arrow will bring the first or last interleave, respectively, to the top of the card. The scroll window in the upper right will let you view different portions of the card.Many programs use dashes to indicate gaps. Note that Aligner assumes that you use periods (".") for gaps, not dashes ("-"), because dashes are used for match characters (as in Eugene). You may need to replace dashes with periods if you have aligned your sequences with another program. DNA Translator (under N-str field menu) or a word processor can be used for this conversion.To change the alignment for a particular sequence, click on the corresponding number in the left margin. If you have imported named strings (DNA Translator format), you can also select a sequence by clicking on its name in the scrolling list. If you want to keep the cursor where it is, try typing a backslash, followed by a click on the sequence to edit. Any of these actions will activate the chosen sequence for editing. Move the cursor to the desired point of entry, click to get the standard "I-Bar" editing cursor, and type away. Normally you will only be entering or deleting gaps (periods), which when entered will sound a harpsicord tone if the speaker button is hilighted. You can also type in valid nucleotide or amino acid symbols, which will be spoken if the speaker button is hilighted. You can also use the sequence speaker to read back a sequence for proofreading. Click on a desired starting point, or start at the beginning if no text insertion point is selected, then choose "Speak" from the Edit menu (or type "s" with the command key depressed). Speed and volume are adjustable with the buttons to the right of the speaker button.Typically, manual sequence alignment involves adjusting all but one or a few sequences. For convenience, you can use option-period to enter a gap to all but the currently selected sequence, i.e., a sequence which has one or more "inserted" base pairs/residues. If you also depress the shift key, you will be able to select the number of gaps to add (default is 2). Now typing comma (",") with the option key depressed will delete all gaps (periods) or question marks one site prior to insertion point. If you also depress the shift key, you will again be able to select the number of sites to delete gaps/question marks from. Make sure that your cursor is where you think it is (i.e., not up or down one interleave) before performing these actions.On Macs that support 256+ color levels, and have plenty of memory allotted to HyperCard (?about 2.2+ MB), a Color menu appears which will allow coloring of nucleotides (top three menu items) or peptides ("Amino Acids" and "AA Properties" menu items). The ability to color-code nucleotide triplets according to their coding ("Codons") is especially useful. Alternative genetic codes are supported as selected from a scrolling dialog box. After you have chosen a code to use, this information is stored directly after each line of the sequence name field (separated from the name by a comma) so that you need select the coding only once. The "Properties…" and "AA Properties" menu items allow display of amino acids according to chemical, functional, hydrophobic/hydrophilic, and ionic charge groups, rather than individually. Choose "First Block" rather than "Color All" to save time by only coloring the first interleave block. The coloring is still rather slow, but it presents an interesting view of one's data. Photographing the screen should be feasible, or use a utility such as the shareware control panel devise Flashit (ftp to 'sumex-aim.stanford.edu' and 'get info-mac/cp/flash-it-22.hqx') to capture the color screen to a PICT file or the clipboard. I have had good results pasting PICT images into a word processor (Nisus) document and then printing the document on a Hewlett Packard Deskwriter C (these cost about $600). If you want to get the color PICT printed on a postscript color printer, you will need to paste the PICT into some program that can support a conversion to standard CMYX mode (e.g., Adobe Photoshop). If you just need a more standard 4-color display of nucleotide base pairs, you may find this feature too slow for convenient use. Another, newer, approach is available by choosing "Color Window 1" or "Color Window 2" from the Color menu. So far, only bases are supported, but amino acid coloring will soon be implemented. As above, text editing is not allowed. Depress the option key while selecting one of these menu items for additional color scheme choices (also see version history for 1.0k below).Under the File menu is an item called "Create Nuc Gaps". This is intended to make it easier to maintain or create a nucleotide alignment which corresponds to a peptide alignment. DNA Translator allows you to extract corresponding protein-encoding sequences from dynamic gene maps (e.g., 9 animal mitochondrial DNAs), which can then be translated to inferred peptide strings (including support for alternative genetic codes). Nucleotide and peptide examples of such output (Cytochrome oxidase b) are included as sample data. If you import the peptide example (use "Import Multiple"), note that it is already aligned, then select "Create Nuc Gaps" and choose the corresponding nucleotide example file (unaligned). This will enter triplets of periods at the proper places in the nucleotide strings, which can be imported to Aligner or output as a text file. Follow this example to do your own conversions, making sure that they are likewise "translator" format files which correspond in their sequence order and in their starting (5') positions.Important note: If you are running MultiFinder and having trouble getting the card window to expand to the size of a large screen screenrect, then you need to increase the size of the MultiFinder partition allocated to HyperCard:1. Quit HyperCard2. Click once on the HyperCard application3. Type 'I' with the CommandKey (Get Info)4. Type in 2,000 or more into the dialog box.[Special note for those with color monitors smaller than the standard 13" (640 x 480 pixels), you may need to experiment some with "Small/Large Window" and "Shorten/Lengthen Card" under the Edit menu in order to color your data correctly.]Version History:.99.8 First posting 9/24/91 (starting version number coincides with that of current DNA Translator stack).99.9 10/1/91. Changed appearance, made "Print Card" a bit more useful, fixed a few minor problems..99.9b 10/3/91. Fixed a one-line script error introduced in .99.9.1.0b 11/24/91. Fixed a problem involving resetting Match/Unmatch menu item, which now resets to the last active setting for a particular card. Also, added an option to select a new sequence for editing by clicking on its name in the scrolling name list.1.0c 11/27/91. Fixed more problems with Match/Unmatch, including bug encountered when exporting multiple sequences in interleaved matched format. Also made it impossible to add spaces while editing sequences, or to edit sequences while sequences are matched. Added the close box to this help file.1.0d 12/13/91. Fixed more problems with Match/Unmatch: As in DNA Translator, now periods (gaps) are not matched and problem with matching on overhangs of variable sequence length fixed. Periods are added to termini of all but the longest sequence until sequence length matches the longest sequence. This was a serious bug which could have corrupted the ends of sequences. Made it possible to enter "?" while the sequence speaker is on. Fixed the "Print Card" command. New feature added to allow selecting a new sequence to edit by typing a back slash ("\"), followed by a mouse click on the new sequence to edit (while the cursor remains as a heavy "plus").1.0e 12/28/91. Fixed more problems with Match/Unmatch: Now does what it was supposed to in 1.0d above. Added a feature to add a gap "." to all but the currently selected sequence whenever "." is typed with the option key depressed. This avoids the necessity to select each sequence to add a gap to all but one sequence.Versions below are copyright 1992 by Douglas J. Eernisse1.0f 1/3/92. Added automatic card number field above navigational arrows in upper left of card.1.0g 1/10/92. Gap feature described in 1.0e fixed to force unmatching before addition of multiple gaps. A new feature was added to permit alignment of nucleotide strings to preserve a peptide alignment. Resulting nucleotide strings can be imported or exported to a file. The sample data was modified so that the nucleotide and peptide data correspond, a necessary requirement for this feature. See instructions above.1.0h 1/20/92. Added handlers to stack script to instruct users who have either too little memory allocated to HyperCard under MultiFinder or insufficient RAM, so that the card window is shown much smaller than a 640 x 480 pixel monitor. Added Color menu and features, made possible by external commands contained in the freeware stack Colorizer 1.0 (see stack script for credits and instructions above). A shorten/ lengthen card size option was added, so that length can be adjusted between 640x1200 and 640x520 pixels. Colorizing requires the creation of two full-card screen buffers, so card size shortening reduces Colorizer-specific RAM requirements from about 1500K to 650K. Users who wish to view more than about 40 sequences will need to lengthen the card size. Doing so will remove the Color menu unless you have allotted plenty of RAM (3000K+) for HyperCard. Memory management probably still needs to be fine-tuned.1.0h(2) 1/25/92. Only fixed problem with "Create Nuc Gaps", and with menu item "Match" or "Unmatch". Both problems were added as the result of other changes, but the former was serious because gaps were introduced in the wrong places.1.0h(3) 1/27/92. Fixed problem with coloring of 4th to nth interleaves. Added short instructions for testing coloring feature as a text file in "Aligner Sample Data" folder.1.0h(5) 2/14/92. Modified "Import Multiple" so that a user can have a return character after the last line of the import file. Also modified importing to recognize whether data is already matched or unmatched, and the "Match/Unmatch" menu automatically changes when appropriate. Careful that you do not try to import data with dashes corresponding to gaps (see above)! Modified "Export Multiple" so that coding information can be optionally preserved. Modified selection of coding to return to the last coding selected when the user is entering multiple codes. Modified coloring to permit coloring of matched data (which is temporarily unmatched). Improved the "New Card" feature and added a "power user" feature enabling the clearing of all data on all cards when "Clear Data" is selected from the menu with the option key depressed. Likewise, selecting "Delete Card" with the option key depressed deletes all but the first two cards (which are protected from deletion).1.0i 3/12/92. Added some more keyboard short-cuts (see above).An enhancement to DNA Translator also affects Aligner. Now you can extract multiple sequences directly to Aligner, either as nucleotides or converted to peptides. Try clicking on an animal mtDNA gene button with the command and shift keys depressed to test this new feature. You will be sent to Aligner with nine, ready-to-align DNA or amino acid sequences for the gene corresponding to the button you clicked on.1.0j 4/13/92. Added an error-checking facility. After performing an alignment it is desirable to check the data against the starting strings. This is done by selecting "Error Check" from the Edit menu. The card's current sequences will then be compared to corresponding ones in a file specified in a prompt dialog, ignoring gaps and strings of missing characters (which may get expanded or contracted during alignment) in both the present alignment and the original string data. The order and number of strings must be identical in each. The user will also be prompted to supply a report file name, in which results will be listed, sequence by sequence. If differences are found, the strings minus any gaps or missing characters will be placed sequentially in the report to aid in locating the discrepancies. Error checking is highly recommended even if you think that you were careful. Ordinarily you would just check your aligned sequences against the file that you imported.A feature was added so that clicking on row numbers while holding the option key down will briefly display the name of the sequence (assuming named strings have been used). A pointer indicator was also added to indicate the currently active sequence (displayed for the first interleave block only).1.0k 6/8/92. An experimental feature was added to enable coloring text with two new custom externals, ColourText and SetFont, by Nigel Perry, also utilizing a special set of fonts supplied as resources within the stack. This still does not permit editing of the alignment. For best results try starting with a relatively small alignment and select "Color Window 1" or "Color Window 2" from the Color menu. This will display a scrolling interleave of your current data, with text coloring corresponding to the different bases. This feature is still not thoroughly tested. If you experience difficulties, restart Aligner and operations should return to normal.Added another experimental coloring feature, this time using an external called "Textoid" by Rinaldi. In the uploaded version, the necessary externals have been removed. If anyone wants to experiment with use of this external, follow the procedure outlined in the Version History file included with this upload (see 6/12/92 comments) to copy the necessary resources into Aligner. This feature can then be accessed by holding down the command key while selecting "Bases" or "Amino Acids" from the Color menu. A new window will be created and the sequence text will (rather slowly) be colored. The advantage of this scheme is that it permits editing of either nucleo- or peptide alignments in color, however, I have experienced problems with attempting to edit colored sequences so this still needs some work. The adjusted alignment would need to be manually copied and transferred to a field or word processor document at present, but "Copy" commands do not appear to work as they should.1.0k(1) 6/15/92. Added support for coloring amino acid (in addition to previously supported nucleotide) data according to the chemical, functional, hydrophobic/hydrophilic, and ionic charge groups, replacing the "Special..." menu item with "Properties..." and "AA Properties..." for nucleotide or amino acid data, respectively.Added new externals by Nigel Perry. This includes a text coloring routine that uses RGB values instead of the standard Mac 8-color schemes as before. Added the options to the Color menu. Holding the option key down will now use the 8-color scheme. Otherwise RGB values are used.</text>
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<text>Ala Arg Asn Asp Cys Gln Glu Gly His Ile A R N D C Q E G H ILeu Lys Met Phe Pro Ser Thr Trp Tyr Val L K M F P S T W Y V</text>
