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Sequence Analysis of Single Polymorphism in the Factor V Gene - page 11 / 15

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Biol  420                                                                                                                                          Fall 2007

2.

Under the Analysis tab, select the Sequencing Wizard.  Within the Sequencing Wizard window, the Modify Single Sample window can be opened by double clicking in the appropriate row of the sample # column.  If the sequencing has already been performed , then in this manner you can edit individual bases.

3.

If the sample was originally sequenced with the incorrect lane order, you can change the lane order by entering the correct order in the Lane Order field.

4.

Lanes are normally found automatically during sequencing.  However, to ensure that your lanes are properly aligned, the Sequencing Settings tab in the Sequencing Wizard can be used to edit the lanes.  To begin editing, click the Edit Lanes button to enter Edit mode.  When the Edit Lanes tab is displayed, the four lanes lines of the sample are displayed on the image in the e-Seq window.  The green squares on each lane line are movable and should be adjusted to be at the center of each lane.  Once done adjusting all of the lanes, then click the Apply button to save any edits.  This must be done to both the seq.700 and seq.800 image.

5.

To sequence a sample, select where you want to start and stop sequencing and then select Sequence Now.

6.

Print your sequence data.  Select Samples on the Report menu to create a report that contains sequence data from one or more sample files of the current run.

a.

Select the sample(s) from the sample list on the left side of the Report window.

b.

Select the report type  as GCG or EMBL from the drop-down list

c.

Set the destination for the report as Printer.

d.

Click OK.

7.

Print the sequence image by selecting Print on the File menu.

Comparing the sequences

When Li-Cor AlignIR software is installed on your computer, sequence data can automatically assembled or aligned using post-sequencing actions in the Sequencing Wizard.  

1.

First you must create Gallery file versions of your e-seq files.  A gallery is just a special type of run file.  In galleries, images are imported rather than created during electrophoresis.

a.

Select Create Gallery on the Analysis menu.

b.

Click Browse and use the file selection window to set the folder to MCB_Buccal_trial1 and the gallery name to MCB_Gallery_3.  Click Save in the File Selection Window.

c.

Click the Image Gallery tab to make sure it is selected.

d.

Click Add to add an image file to the gallery.

e.

In the MCB_Buccal_trial1 folder, select your MCB_Buccal_trial1700.tif and click Open.

f.

Click Add in the New Gallery window and add your MCB_Buccal _trial1800.tif  image in the MCB_Buccal_trial1 folder.

g.

Make sure the Copy Images …. check box is selected.

h.

Click OK to create the gallery.

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