genome alignment errors with Bowtie2

#1

Hi everyone,

I am new to using Galaxy, and bioinformatics in general. I am trying to align a hummingbird genome to the Anna’s Hummingbird reference genome…I’ve successfully uploaded the fastq and fa files, but the bowtie2 runs fail. I want to produce a bam file that I can combine with my other individuals who have been aligned to the same genome. I’d appreciate any help I can get!

A screenshot of my settings is attached.

The errors I get are:
error
An error occurred with this dataset:
Settings:
Output files: “genome.*.bt2”
Line rate: 6 (line is 64 bytes)
Lines per side: 1 (side is 64 bytes)
Offset rate: 4 (one in 16)
FTable chars: 10
Strings: unpacked
Max bucket size: default
Max bucket size, sqrt multiplier: default

and

3 Bowtie2 on data 2: aligned reads (BAM)

error

An error occurred with this dataset:

Settings: Output files: “genome.*.bt2” Line rate: 6 (line is 64 bytes) Lines per side: 1 (side is 64 bytes) Offset rate: 4 (one in 16) FTable chars: 10 Strings: unpacked Max bucket size: default Max bucket size, sqrt multiplier: default

28%20AM
Screen Shot 2019-08-12 at 10.37.28 AM.png2498×2366 468 KB

#2

Hello,
From your screenshot it looks like you are using the fasta file as the (reads) input and mapping to a Baboon reference genome. Try providing your reads to the “FASTA/Q file” input field and select the ‘use from history option’ to use the fasta reference as the reference genome.

Give that a try and let us know if issues persist.

Thanks for using Galaxy!

Mo Heydarian

#3

Thanks for the reply. I re-ran it, and the run finished, however, the bam and bai files were only 2.5 kb in size each and contain gibberish.

I’m using the original fastq file that I converted from the original raw reads from ncbi (Anna’s Hummingbird 17kb cut on Blue Pippin and 110 pM loading concentration 10 PACBIO_SMRT (PacBio RS II) runs: 1.6M spots, 26.3G bases, 87.5Gb downloads, Accession: SRX1131887) using SRA and the .fa file I downloaded from ncbi as well. My actual samples are aligned to this .fa file, and I need Anna’s as an outgroup, so I need a .bam file to include with the .bam files I have that are aligned to the same genome. I just thought I’d include all the background in case there’s something else I’m doing wrong.

Command line from Galaxy: @PG ID:bowtie2 PN:bowtie2 VN:2.3.4.1 CL:"/cvmfs/main.galaxyproject.org/deps/_conda/envs/mulled-v1-7576289d51ff5aefa21c8a2af901e1a61eb245f4e5fd66ae894d120dd35ff8f6/bin/bowtie2-align-s --wrapper basic-0 -p 6 -x genome -U input_f.fastq"

I’d really appreciate it if you could provide some assistance.

Thanks,

Brian