Question

FastQ files from MiSeq Illumina to the "mapping file" in Qiime...Barcodes sequences needed?

1

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8.5 years ago

chelseabarreto22 ▴ 10

Hello!

I am very new to Qiime and can really use some help for more of the simple stuff!

I am working with soil microbial community data where I have received sequences from Genewiz (Illumina miseq 16S sequencing V3-V5). I am trying to make the mapping file but I don't seem to have the barcode sequences. I believe this data has already been demultiplexed. Do i need the bar code and primer sequence to set up the mapping file or can I do without it? Struggling very hard! I know this is a simple question but any help in getting the fastq files formatted correctly would be appreciated! Thank you!

Qiime Barcode fastq • 5.8k views

ADD COMMENT • link updated 8.5 years ago by pescadordigital ▴ 10 • written 8.5 years ago by chelseabarreto22 ▴ 10

score 1 · Answer 1 · 2016-05-18

Hi,

I already battled with this problem too... until I found the answer. Of course I assume that your FASTQ files were already demultiplexed, ando also assume that you are attempting to use the [split_libraries_fastq] command ( see http://qiime.org/scripts/split_libraries_fastq.html ) command. I also assume that you already merged your paired-end reads file into just one. In the QIIME documentation for this particular command: see [http://qiime.org/scripts/split_libraries_fastq.html] you'll see that there is an option --barcode_type "not-barcoded" , you can use this one.

Putting things together, the QIIME command I used was:

 split_libraries_fastq.py \
    -i $FASTQ\
    -o slout\
    --barcode_type 'not-barcoded'\
    -m $MAP_FILE\
    --sample_ids sample.1\

$FASTQ is the original FASTQ file and $MAP_FILE is your map file.

I fought with the problems to have to analyze multiple FASTQ files and in order to not have the need to write manually a map file each time I needed to run with a new FASTQ file I wrote the following Python code to generate for me a new "dummy" map file each time I need to run QIIME:

import os, sys
from os import path

def main():
    fastq_filename = sys.argv[1]
    if len(sys.argv) > 2:
        new_mapping_file_name = sys.argv[2]
    else:
        new_mapping_file_name = ".".join(fastq_filename.split(".")[:-1]) + ".map.txt" 

    mapping_file_header = ['#SampleID','BarcodeSequence','LinkerPrimerSequence','Description','File']
    mapping_file_line1 = ['sample.1','','','single_file',path.basename(fastq_filename)]

    with open(new_mapping_file_name,"w") as f:
        f.write("\t".join(mapping_file_header) + "\n")
        f.write("\t".join(mapping_file_line1) + "\n")


    print "New mapping file is ", new_mapping_file_name

if __name__ == '__main__':
    main()

This script accept two arguments: the first being your FASTQ filename and the second the name you wish to give the map file. As you can see in the code the map file is a tab-separated values file with just two lines: one is for the headers and the second has empty contents for the 2nd and 3rd columns. 'sample.1' is a dummy name I chose to give to the sample you can modify it at will: QIIME won't bother with that I think but if you change it, just remember to change the sample name accordingly in the command line option -sample_ids.

Cheers