BCHE 451/MOLB 550
Feb. 18, 1998
Importing and Viewing Chromatographic Trace
Files in Sequencher 3.0
- Use an FTP utility to connect to pc-plammers.nmsu.edu using the login
ID and password given out in class
- Select Binary transfer
- change directories to "data". This directory contains real
sequence data from LI-COR machines in NMSU's DNA sequencing facility
- open a directory from Feb. 98 like 020598A9
- choose at least 4 files with the .SCF suffix. These are chromatographic
trace files and are 30 to 60 K in size
- Try to pick sets of two .SCF files with the same name but different
primers eg: T7 and T3 or For and Rev.
- Go for the biggest files to maximize the chance of overlapping sequence
from the two ends
- Get those files (place in your Sequencher folder on the HD or on your
floppy drive for future use)
- Open Sequencher application
- Select Finder from the applications menu (TOP right-hand corner of
monitor)
- Use the Macintosh "Drag and Drop" utility to import all the
.SCF files into Sequencher (Drag the .SCF icon on to the open Sequencher
icon)
- You should see a new icons for each .SCF file appear in the Project
Window of Sequencher.
- Double click one icon to see the sequence
- Go to Window -> Chromatogram to display the chromatogram
- What do the double lines of assigments mean?
- Go to User Preferences and select Chromtgrm, the play with the features
there to see how the window display changes
- Stack the Sequence window under the Chromatogram window (set with just
one line of trace info) so both can be viewed on the monitor simulaneously.
- Highlight a base in either window. What happens?
- Find some regions where the base called doesn't seem right to you...(maybe
down near the end of the file)
- Make the change you think should be made
- How does Sequencher keep track of your edits?
- Do an assembly on your four DNA's to see if you have a region that
is represented on both strands
- If you can find a contig: Bring up the contig assembly window
- Select a contig column by clicking on its base in the consensus line
- Go to Window -> Chromatogram to and the chromatogram for that region
of the contig should appear!
- Do some more editing
- Save all work that you do as part of this exercise because it will
be part of the project report