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Old 08-21-2013, 01:15 PM   #10
GenoMax
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Location: East Coast USA

Join Date: Feb 2008
Posts: 7,077
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It sounds like even though your files were not demultiplexed they were somehow sorted on the tags so that all the corresponding R1 and R3 (based on R2) were in the same order in the original files. If you have managed to separate the samples into 6 files are you able to write a script that will add the "tag" to the ID's of the separated files so that you end up with something that looks like below (see changes marked in red):

R1 file
@IPAR1:2:1:4029:1196:1#NNNNN/1

R2 file
@IPAR1:2:1:4029:1196:1#NNNNN/2

If you are not able to do this yourself then the better option is below

The script included in "qiime" package will take as input the R1 file along with the R2 (tag file) and then will produce separate files for each of your samples (sounds like you have 6). You will then repeat the process with R3 file along with the R2 (tag) file to produce the corresponding files that will contain the paired-end reads.

You can run the qiime script as follows (Disclaimer: I have not used qiime script myself but based on the info provided on the help page I expect the script to work as noted below).

Code:
$ split_libraries_fastq.py -i /path_to/Read1.fastq -b /path_to/Read2.fastq --store_demultiplexed_fastq --barcode-type 8 --sample-id replace_with_sample_name -o output_dir_name
Repeat for Paired-read
Code:
$ split_libraries_fastq.py -i /path_to/Read3.fastq -b /path_to/Read2.fastq --store_demultiplexed_fastq --barcode-type 8 --sample-id replace_with_sample_name -o output_dir_name

Last edited by GenoMax; 08-21-2013 at 01:53 PM.
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