Question: HISAT2 produces no aligned reads
0
gravatar for srinivk
6 months ago by
srinivk0
srinivk0 wrote:

Hello all,

My lab has just installed HISAT2 on our instance of Galaxy. Unfortunately I am having some trouble. When I run HISAT2 (input was fastq files with metadata as fastqsanger), the job completes however the resulting BAM file is only ~ 2 kB and when viewed in Tablet there are not reads. HISAT2 did not give any error messages either. Does anyone know what is going on, and what are possible solutions?

Thanks

Edit: Some runs show this message

gzip: input_f.fastq.gz: not in gzip format gzip: input_r.fastq.gz: not in gzip format 0 reads 0.00% overall alignment rate

while others show

Building DifferenceCoverSample Building sPrime Building sPrimeOrder V-Sorting samples V-Sorting samples time: 00:02:06 Allocating rank array Ranking v-sort output Ranking v-sort output time: 00:00:26 Invoking Larsson-Sadakane on ranks I

rna-seq • 319 views
ADD COMMENTlink modified 6 months ago by Jennifer Hillman Jackson23k • written 6 months ago by srinivk0
0
gravatar for Jennifer Hillman Jackson
6 months ago by
United States
Jennifer Hillman Jackson23k wrote:

Hello,

Start by checking the actual format of the data versus the assigned datatype. It looks like you have an uncompressed fastqsanger datatype assigned to describe what is actually fastqsanger.gz compressed data input.

How-to: https://galaxyproject.org/support/compressed-fastq/

All support topics linked here: https://galaxyproject.org/support/ These cover the most commonly seen usage issues along with methods to resolve problems, including the one above.

Check, make changes, and rerun. Let us know how it goes if you still have problems.

Thanks! Jen, Galaxy team

ADD COMMENTlink written 6 months ago by Jennifer Hillman Jackson23k

Hi Jennifer,

I looked into the file format. We have confirmed these are uncompressed fastq files (ranging from 5GB to 28GB). Looking through the code, we suspect that HISAT may be thinking our inputs are compressed hence the input_f.fastq.gz and input_r.fastq.gz assignments.

This is the command that runs

ln -f -s '/galaxylab/production/new/galaxy/database/files/011/dataset_11880.dat' input_f.fastq.gz && ln -f -s '/galaxylab/production/new/galaxy/database/files/011/dataset_11897.dat' input_r.fastq.gz && hisat2 -p ${GALAXY_SLOTS:-1} -x '/galaxylab/production/new/galaxy/tool-data/hg38/hisat2_index/hg38/hg38' -1 'input_f.fastq.gz' -2 'input_r.fastq.gz' -k 5 --max-seeds 5 | samtools sort - -@ ${GALAXY_SLOTS:-1} -l 6 -o '/galaxylab/production/new/galaxy/database/files/013/dataset_13223.dat'

Any furthur help would be appreciated. Thanks.

ADD REPLYlink modified 6 months ago • written 6 months ago by srinivk0
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