Generating SFF files

[Xterra]

By mistake the SFF file was deleted once the Amplicon Full processing pipeline was done. How can we reprocess the data to generate the SFF file back again?
Thanks!

[kmcarr]

By mistake the SFF file was deleted once the Amplicon Full processing pipeline was done. How can we reprocess the data to generate the SFF file back again?
Thanks!

Sadly the gsReporter application will not produce SFF files from an exiting analysis directory; SFF files can only be generated by gsRunProcessor. But you don't have to repeat the entire Signal Processing pipeline, you can perform an Analysis Filter job on the existing D_ directory which will take far less time.

If you are unfamiliar with how to perform a Filter only analysis (these steps assume your current working directory to be the run (R_) directory and the Full Amplicon analysis directory (D_) still exists there):

1. Create a template file with the default amplicon filter settings, saving it to a file named (e.g.) ampliconFilter.xml:

# gsRunProcessor --template=filterOnlyAmplicon > ampliconFilter.xml

2. Run the filter only analysis pipeline providing the name of the template file you just generated and the existing analysis (D_) direcotry.

# runAnalysisFilter --pipe=./ampliconFilter.xml <D_directory_name>

The filter only pipeline runs much faster than the full analysis pipeline. You will have a new, complete D_ directory with the SFF files.

[Xterra]

kmcarr

I actually did exactly what you suggested but the new D directory did not included the SFF file on the regions subfolder.

[kmcarr]

I actually did exactly what you suggested but the new D directory did not included the SFF file on the regions subfolder.

The SFF files are not in the regions subfolder. There should be a separate subdirectory named 'sff'.

[Xterra]

kmcarr

Sorry that's what I meant. Now, I am not quite sure what's going on, I repeated the process three time and I finally got the SFF file. However, when I split the MID, I can see I got very few reads/MID. When I split the original SFF file that came out of the run, I had plenty of reads per MID. Therefore, something is wrong.
Now, I can see the fasta and qual files, can I split the MID from the fasta file using any of the SFF tools?

[sklages]

Sorry that's what I meant. Now, I am not quite sure what's going on, I repeated the process three time and I finally got the SFF file. However, when I split the MID, I can see I got very few reads/MID. When I split the original SFF file that came out of the run, I had plenty of reads per MID. Therefore, something is wrong.
Now, I can see the fasta and qual files, can I split the MID from the fasta file using any of the SFF tools?

Why do want to do that? You should investigate on what is going wrong with your filtering job ...

just my 2p,
Sven

[Xterra]

At this point I am more interested on retrieving my data.
Do you know of any tool I can use to split the MID using the central FASTA file that was generated during the run?

[sklages]

You finally will need to find out what was wrong, believe me ... :-)

Nevertheless, one of the tools capable of doing what you want might be "FASTX Barcode Splitter" from the fastx toolkit (http://hannonlab.cshl.edu/fastx_tool...splitter_usage).

hth, Sven

[Xterra]

sklages

Thanks! I will check it out.