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I wrote a script to generate project files for SeqTrace
and perform additional trim operations using a reference
sequence.  To complete my needs it would be great if SeqTrace
could process project files without the GUI interface. 

Having read through the source code of SeqTrace, it does
not seem trivial to add this functionality. 

Separating the GUI from the calculations required would also 
make this code more useful over the long term. 

Any tips on how to proceed with this work would be much 
appreciated. 

Regards,
David  

What steps will reproduce the problem?

I have forward and reverse reads but the finished sequence is not being 
created. The sequences are aligning fine but there is no "finished sequence". I 
assume this is happening because there are no confidence scores. Apparently the 
chromatogram trace files do not include the scores, or they are no showing up. 
(I have attached an image)

Is it still possible to create a finished sequence? I tried to set the minimum 
confidence to 0, but that did not fix it. The program would freeze when 
generating the sequence.

What version of the product are you using? On what operating system?
Seq Trace 0.8, Windows 7.

Thanks. 

The .abi files given to me by my sequencing core do not contain quality data 
about the base calls. Consequently, seqtrace is reading the confidence of every 
call as '0', and will not let me edit the files or do anything because it 
requires a value for minimum confidence. If this could be changed to have an 
option to ignore quality data and allow me to still work with the sequences, 
that would be awesome. I like this program, but unfortunately I'm looking for 
alternatives because of this issue.

Tested on lubuntu 14.04 and windows 7 x64 with the latest version as of this 
post. 

Thanks.

Hello,

  I am running SeqTrace 0.9.0 using Ubuntu Linux v 13.1.0. I am using SeqTrace to view and edit ABI sequencer chromatogram files. However, every time I try to edit sequences, the menu buttons at the top of the sequence editing window are grayed out as are the editing options under the 'Edit' pull down menu. This happens when I open single or batch chromatogram files.  

  The sequencer machine used is an ABI 3130XL running KB Basecaller software v 1.4.0.

Any help you can provide will be greatly appreciated!

Thanks,
George

What steps will reproduce the problem?
1. Create New Project and Choose Location of Sequencing Files
2. Add Trace Files and then Add Trace Files to Project
3. Select Trace Files and then Generate Finished Sequences for All Trace Files

What is the expected output? What do you see instead?
The expected output is a Sequence Alignment of Forward and Reverse Reads that 
will show consensus alignment
Instead, I see a box that says, "Generating sequences for all trace files... 
0%"  and it stays on this until I quit the program.
What version of the product are you using? On what operating system?
Python 2.7; Seqtrace    OSX 10.9.6 Mavericks

Please provide any additional information below.

When I first downloaded and used Seqtrace (0.9.0) I had Kubuntu 12.04 and 
everything worked just fine. So I updated it to Kubuntu 14.04 and since then 
the numbers inside text boxes in "Project settings" are invisible (see attached 
image). It happens whenever I am starting a new project or opening an old one. 
On either case the default settings, my old project settings, or newly created 
project settings appear to be there due the behavior of the consensus 
generation, but the fact that the numbers won't show up next time I open the 
dialog remains annoying.

I supposed the issue could be due the update, some confusion with the python 
packages, but recently I have installed a new, fresh-formatted Kubuntu 14.04 
and the problem is still here. I'm guessing it's some incompatibility with a 
new version of some or any of the pythons packages in the Ubuntu repository?

Thank you in advance.

When I first downloaded and used Seqtrace (0.9.0) I had Kubuntu 12.04 and 
everything worked just fine. So I updated it to Kubuntu 14.04 and since then 
the numbers inside text boxes in "Project settings" are invisible (see attached 
image). It happens whenever I am starting a new project or opening an old one. 
On either case the default settings, my old project settings, or newly created 
project settings appear to be there due the behavior of the consensus 
generation, but the fact that the numbers won't show up next time I open the 
dialog remains annoying.

I supposed the issue could be due the update, some confusion with the python 
packages, but recently I have installed a new, fresh-formatted Kubuntu 14.04 
and the problem is still here. I'm guessing it's some incompatibility with a 
new version of some or any of the pythons packages in the Ubuntu repository?

Thank you in advance.

Since the alignment does not take into account quality data, it causes some 
final sequence errors which would logically be ignored during manual 
inspection. All settings were following the default install except minimum 
quality which was set to 20 for the purpose of showing Example 2. 

Example 1: Insertion errors (insertions.png)
Trace 1 has a high-quality trace which says CC. Trace 2 is just beginning, with 
a low quality N added into the sequence. This results in a final base call of 
CNC which is clearly not the case. 
Example 2: Bayesian poisoning due to misalignments (starting.png)
Trace 1 has a low-quality starting trace, which is misaligned. It has a C with 
a quality of 23. The misalignment pairs it with a G with a quality of 28, which 
is marked as N due to the disagreement, throwing off the Bayesian base caller. 
Previous bases (the A and C) with lower qualities are called correctly. 

To suppress errors of the first kind, code might be added to look for 
insertions of N within a high-quality (above minimum threshold) and 
automatically remove these insertions. 
To suppress errors of the second kind, it could be possible to implement a 
"trace-trimming" feature, using the same code used to trim the final sequence, 
in order to remove misaligned starts and ends of traces. 
Sinces traces also suffer from clusters (~10 bases) of low quality data points 
from 20-160 bases, they should also be appropriately treated when it comes to 
alignments. 

The perfect solution would be to have an alignment algorithm which takes 
quality into account, but lacking those, these aforementioned things will be 
good stopgaps. 

What steps will reproduce the problem?

1. Once the final alignment is made between forward and reverse trace files, 
the y-scale bar no longer is adjustable. If the y-scale bar remained functional 
it would be helpful. 
2.
3.

What is the expected output? What do you see instead?

The y-adjust bar is always stuck in the final view of forward and reverse 
together.  In most instances the trace file is scaled acceptably so that I can 
see both forward and reverse peaks to judge each base call.  However, in some 
sequences, once the forward and reverse are put together, the y-bar is turned 
all the way down, and the peaks are too low to read.  If I could raise the 
peaks it would be helpful. 


What version of the product are you using? On what operating system?
Version 0.8.1 for windows

Please provide any additional information below.

The bar works fine when an individual trace (only forward or only reverse) is 
being viewed.  It is only stuck when the two are put together. 

I have attached the two trace files (ITS1 is forward, ITS4 is reverse) that I 
had the problem with in which the peaks were too low.  However, note that the 
y-adjust gets stuck in all files, not just these.  It became a problem with 
these because due to the low y-scale the peaks are too low to read.