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

For instance if you click the Clean Data radio button, contig assembly is faster but
Sequencher may miss some possible matches if the ends of your sequences contain a
large number of ambiguities. For best performance, make sure that you have trimmed
poor quality data from your sequences.
The Dirty Data algorithm is somewhat slower because Sequencher is performing more
rigorous comparisons between the sequences. You should note that automated sequencers
create dirty data and Sequencher’s algorithm is optimized to deal with that.
The Large Gap algorithm allows you to assemble sequences that are expected to contain
insertions and deletions (gaps) greater than 10 bases long. Such gaps are typically found
in evolutionary studies and sequence comparisons from different organisms. The Large
Gap algorithm is similar to and slower than the dirty data algorithm but allows larger
gaps to occur when performing the assembly. Typical examples of assemblies that require
the Large Gap algorithm include the comparisons of a cDNA sequence to a genomic
sequence and the assembly of related genes with alternative splicing.
Assembling with Dirty Data
Go to the Assembly Parameters dialog box and select the Dirty Data radio button.
There are two sliders for changing minimum match values. To set the proportion of bases
that have to match use the Minimum Match Percentage slider. To set the minimum
number of bases that must overlap use the Minimum Overlap slider.
Change the settings by positioning your cursor on the slider, holding the mouse button
down and dragging the slider left or right. The value set will automatically update as you
move the slider.
For example, if you attempt to assemble a 17-base sequence that matches perfectly to a
selected 1000 base sequence with the minimum overlap set at 20 bases (Sequencher’s
default minimum overlap), the two fragments will not assemble. You must first reduce
minimum overlap to 17 bases or lower and then assemble the fragments.
Assembling with Clean Data
When you select the Clean Data algorithm, an additional option called Maximum Loop
Out Size is available. This has an elevator button for adjusting size value (Figure 9-2
below). This parameter is the largest number of consecutively mismatched bases that are
acceptable in a potential overlap. Change the setting by positioning your cursor on the
appropriate arrowhead, and holding the mouse button down. The maximum value, which
can be set, is 6.
To set the proportion of bases that have to match use the Minimum Match Percentage
slider. To set the minimum number of bases that must overlap use the Minimum Overlap
slider.
SEQUENCHER 4.8 User Manual for Windows © 1991 - 2007 Gene Codes Corporation, Inc. All rights reserved.
65