Sequencher 4.8 User Manual--PC - Bioinformatics and Biological ...

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Setting Vector Trimming Criteria Sequencher has a set of default criteria to facilitate the recognition of any contaminating sequence. These criteria are listed in the top region of the Vector Insertion Sites window. If these parameters do not work for your specific circumstances, you can alter the vector screening parameters. As an example, if there is ambiguity in the region of the vector junction, you can increase the likelihood that Sequencher will recognize the vector sequence by decreasing the approximate match percentage. If it is essential to remove even a single base of vector contamination, you can decrease the minimum overlap to be considered as vector contamination. You can change any of the five independent parameters in order to improve your trim. Elevator buttons are used to increase or decrease the parameter value. The following table describes the recognition criteria for contaminating sequence. Parameter Minimum overlap to consider as vector contamination Approx. match percentage to consider as contamination Minimum overlap allowed without exact matches Additional bases to remove from a contaminated 5’ end Additional bases to remove from a contaminated 3’ Description The smallest amount of overlap to consider as vector contamination. The default value is three bases. The percentage match Sequencher needs to find before considering the matching sequence to be vector contamination. Ambiguities do not count as full matches or mismatches. The smallest number of matching bases within which an inexact match is permitted. If fewer bases than this match, they must match exactly. Extra material, if any, to be trimmed off the 5’ end of the remaining DNA sequence along with the vector contamination. The default is zero bases. Extra material, if any, to be dropped off the 3’ end of the remaining DNA sequence along with the vector contamination. The default is zero bases. Once you have finished choosing vectors and setting recognition criteria close the Vector Insertion Sites window. SEQUENCHER 4.8 User Manual for Windows © 1991 - 2007 Gene Codes Corporation, Inc. All rights reserved. 40
How Sequencher screens sequences When you execute the Trim Vector command, Sequencher screens the selected sequences for vector contaminants using the vector sequences you specified. Sequencher starts at the designated insertion site, the bullet, and then scans outwards. Any sequences that are found to contain vector bases are collected in the Vector Contamination window. (See Figure 6-9.) Please note that any contigs you have selected will be ignored, as will any sequences containing fewer than 21 bases. Figure 6-9 The Vector Contamination window You can print out the Vector Contamination window as a record for your lab notebook by going to the File menu and choosing Print. This is particularly useful if you use the Show Bases button to show which sequences are to be trimmed. You can also sort the window using various criteria by clicking on Sort Items. Executing the trim command To perform the vector screen and trim, go to the Sequence menu and choose Trim Vector…. Once you have reviewed your data and are happy with the criteria you have set, proceed with the trim by clicking on the Trim Checked Items button. Sequencher warns you that this command cannot be reversed with the Undo command. To proceed, click the Trim button. Any red lines or bases will disappear from the Ends Trimming window. Now you can dismiss the window with the command from the File menu called Close Window. If you wish to revert some or all of your sequences to their original state, you can go to the Sequence menu and click on the Revert to Experimental Data command. Remember to save your work at this point by going to the File menu and using the Save Project command, since none of your trim work will have been recorded onto disk. Alternatively use the Auto-Save function to save your work at user defined intervals (see Chapter 21 “Customizing Sequencher and User Preferences”). SEQUENCHER 4.8 User Manual for Windows © 1991 - 2007 Gene Codes Corporation, Inc. All rights reserved. 41
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THIS IS AN AGREEMENT BETWEEN YOU, T
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© 1991 - 2007 Gene Codes Corporati
Page 6 and 7: Templates .........................
Page 8 and 9: Set Circular Genome Size ..........
Page 10 and 11: The Bases View ....................
Page 12 and 13: Protein translations for the Consen
Page 14 and 15: Setting header, footer and margin o
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Page 18 and 19: We welcome your comments or support
Page 20 and 21: Navigating Sequencher Help The Sequ
Page 22 and 23: Contig menu The commands in the Con
Page 24 and 25: Page Intentionally Left Blank SEQUE
Page 26 and 27: Figure 3-1 An empty Sequencher Proj
Page 28 and 29: Figure 3-4 Project Window with Larg
Page 30 and 31: value in the Confidence pane of the
Page 32 and 33: Editing the information for a seque
Page 36 and 37: How to import using menu commands I
Page 38 and 39: Figure 4-2 Browse for Folder dialog
Page 40 and 41: Importing Confidence Scores Many ba
Page 42 and 43: If you have finished working with S
Page 44 and 45: Selecting sequences Selecting seque
Page 46 and 47: Renaming a sequence or contig Renam
Page 48 and 49: Closing the Project Window When you
Page 50 and 51: Performing a trim with default sett
Page 52 and 53: additional control within the Trim
Page 54 and 55: Figure 6-7 The polylinker window No
Page 60 and 61: Basic Reference Sequence properties
Page 62 and 63: When the To Reference by Name comma
Page 66 and 67: A locked editor is read-only. You c
Page 68 and 69: If your sequence was called “MyPU
Page 70 and 71: Viewing summary information in the
Page 72 and 73: Figure 8-7 ORF selected by Next Met
Page 74 and 75: If you want to assign a personal an
Page 76 and 77: Voice verification Sequencher provi
Page 80 and 81: Chapter 9 “Sequence Assembly” f
Page 82 and 83: Figure 9-2 The Clean Data algorithm
Page 84 and 85: Performing the assembly Automatic a
Page 86 and 87: Figure 9-5 Interactive assembly sho
Page 88 and 89: The new contig will appear in the P
Page 90 and 91: When you use a Name Delimiter, it w
Page 92 and 93: If the Expression is a delimiter bo
Page 94 and 95: Figure 10-6 The Project Window with
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Page 100 and 101: Figure 11-2 Contig overview Fifure
Page 102 and 103: Base Numbers At Transitions Clickin
Page 104 and 105: Label Name Function E F G H Sequenc
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Consensus to Forensic Standards In
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consensus is from a base with a low
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move the gaps to the 5’ end, sele
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Figure 12-4 Fill Void dialog box De
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First select the base or range of b
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Page Intentionally Left Blank SEQUE
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The Translated Variance Table If yo
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Structure of the Translated Varianc
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(see Figure 13-5). It is possible f
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Figure 13-7 Translated Variance Tab
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If you want to restrict the Compari
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Figure 13-13 Creating a Joined feat
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In either case, the method you choo
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You can edit your data directly in
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Figure 13-17 The Translated Varianc
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Figure 13-19 Extract from Variance
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The Population Table lists the vari
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If you want to remove sample sequen
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Select Individual Variance Reports
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If you prefer to copy your Translat
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You can copy your Translated Varian
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Summary report display options Sequ
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Enabling the protein display To ena
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Note: Remember that even if a seque
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If you select the Contig Chromatogr
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Select command Table 15-1 Summary o
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Note: The Extend Selection command
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Figure 16-2 Restriction map options
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example, if you were displaying onl
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Figure 16-8 Checkmarked enzyme list
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display this view by going to the V
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The default setting shows exact mat
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Creating and editing features To cr
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Quick feature creation To create a
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Figure 18-1 A dimmed icon Finding t
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Note: If you pause between characte
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a dialog box (Figure 18-6). This le
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Figure 19-1 Export Options dialog b
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You should remember to provide a re
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Figure 19-3 Copy As Protein Transla
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[Project Name] and [User Name]. You
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Colors as Backgrounds Sometimes, di
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Choosing a genetic code You will fi
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time you select the Review button f
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Label & Name The Label & Name prefe
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Click the Use Blank Project radio b
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Use the Columns For Multiple Traces
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If you are making changes to the wa
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Variance Table The Variance Table p
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Figure 22-1 The Validate mtDNA Prof
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Figure 22-3 The Circular Genome Siz
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Menu/Window Command Windows Select
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You may require a more complex deli
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Code Base(s) Meaning 5 A or Another
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Feature Display Color Style Second
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eferences numbers, sequences, or us
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Qualifier type Description Citation
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Feature Key Default Feature Qualifi
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Name CursorProject Window Dialog bo
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