bowtie2 multiple single and paired-end files

[NatashaPavlovikj]

Hi everyone,

I a newbie in Bowtie2, so I have a question about bowtie2 commands for single and paired-end reads.

I have three paired .fastq files, and two single .fastq files.
When I execute the paired files, I use:
bowtie2 -x index -1 file1.fastq -2 file2.fastq -3 file3.fastq ...
When I execute the paired files, I use:
bowtie2 -x index -U file4.fastq file5.fastq ...

The paired files have different number of reads and lines.
These commands are still running, so I wonder did I use them in a right way?
If not, I would appreciate if you could guide me how to use multiple single and paired-end reads.

Thank you very much,
Natasha

[mastal]

bowtie2 multiple single and paired-end files

Hi Natasha,


I think when you have more than 1 file of each type you should separate them with commas, not spaces, and the -3 flag means something related to the index for paired end reads.

so something like this:

paired reads:
-1 filea_read1.fastq,fileb_read1.fastq -2 filea_read2.fastq,fileb_read2.fastq

and for unpaired reads:
-U filec.fastq,filed.fastq,filee.fastq

see the Bowtie2 and bowtie manuals.

Hope this helps,
Maria

[NatashaPavlovikj]

Hi,

Maria, thank you so much for your reply.
You are right that files should be separated by comma.
But, I am still confused about the paired reads.

The three paired Illumina files I have are:
3 ILLUMINA (Illumina Genome Analyzer IIx) runs: 9.5M spots, 2.7G bases

So, my question now is:
should I run these files separately (3 bowtie2 commands, only -1 mate):
bowtie2 -x index -1 file1.fastq ...
bowtie2 -x index -1 file2.fastq ...
bowtie2 -x index -1 file3.fastq ...
or should I run them in a same command (1 bowtie2 command, only -1 mate, separated by commas):
bowtie2 -x index -1 file1.fastq,file2.fastq,file3.fastq ...

Thank you very much,
Natasha

[mastal]

bowtie2 multiple single and paired-end files

Hi Natasha,

If you have 3 files for paired-end reads from Illumina GA, one of the files is probably an index read, so you would have to demultplex the reads in the other 2 files based on the index, before attempting to align the reads, if this hasn't been done already. Check with whoever ran the samples, and have a look at the Illumina website

http://www.illumina.com

for manuals related to the version of the Illumina technology used to generate your reads.

How many samples are in the 3 paired-end files?

As for running paired-end reads with bowtie, you use the flag -1 before the name of the file containing the first reads of the pair, and the flag -2 before the name of the file containing the second reads of the pair.

bowtie2 (other parameters) -1 read1.fastq -2 read2.fastq

see the bowtie manual

http://bowtie-bio.sourceforge.net/index.shtml

best wishes,
Maria

[NatashaPavlovikj]

Maria, thank you very much for your help.
After few tries, I think I have the right scripts now.

Regards,
Natasha

[NatashaPavlovikj]

Illumina put mate 1 and mate 2 in a same file, and when using SraToolkit to convert .sra files to .fastq files, for the paired end files the following command should be executed:
fastq-dump --split-3 file.sra.
That is how the mate 1 and 2 will be created.
This is the answer for my question

Thank you

[sagarutturkar]

Mapping paired-end and Mate-pair reads together

Hi,


I was wondering, How to use Bowtie2 to map paired-end and mate-pair reads simultaneously.

I saw in manual that there are --fr option for PE reads and --rf option for MP reads. But it did not mentioned about using them together.

So if I specify,

bowtie2 -x CLC -a --no-mixed --fr -1 frags.A.fastq -2 frags.B.fastq --rf -1 jumps.A.fastq -2 jumps.B.fastq -S CLC_mapping.sam

would that work?

Thanks
Sagar

[dpryan]

I'm pretty sure that won't work. Your best option, depending on what your goals are, might be to map the two libraries separately and then merge the resulting SAM/BAM files.