Closed kvanraay closed 6 years ago
Update: I put breseq/bin into my $PATH and ran
breseq -r NC_012967.gbk SRR030257_1.fastq SRR030257_2.fastq
I'm running the test_drive data. It's taking a while, but it seems to be working -the first empty READ FILE from my previous run is no longer there.
OK, so I've fixed my problem, which appears to have stemmed from an improper installation. Running the test data just fine now. The main takeaways as a terminal/bash noob:
After unzipping the breseq download, move to the directory where both R and bowtie are (in my case, this was
/usr/local/bin/miniconda3/bin
then pointed to the breseq program using
export PATH=PATH$:/usr/local/bin/miniconda3/bin/breseq-0.33.0-MacOSX-10.9+/bin {my awful $PATH here}
you can find your awful path by typing
echo $PATH
in your terminal.
Glad you solved it!
I think the error in your initial command could have been from not having an output path option flag -o
before the directory /Users/IamUnicorn/Documents/Grad\ School/Projects/Emulsion/IlluminaSequences/breseq/test_drive/
. breseq interpreted that directory as an (empty) input FASTQ file since it it didn't have an option designator in front of it. I'll have to investigate making it give a better error message for that case.
Hello,
I am getting the same error message, but unfortunately none of the advice posted here so far has fixed it. Here is my command:
-> breseq -r Pseudomonas_aeruginosa_UCBPP-PA14_genome.gbk PA14_dphz12_SRR11561537.1.fastq
and the output:
---> bowtie2 :: version 2.4.5 [/Users/john/opt/anaconda3/envs/breseq/bin/bowtie2]
---> R :: version 3.6.3 [/Users/john/opt/anaconda3/envs/breseq/bin/R]
+++ NOW PROCESSING Read and reference sequence file input
READ FILE::PA14_dphz12_SRR11561537.1
Converting/filtering FASTQ file...
Original base quality format: SANGER New format: SANGER
Original reads: 2911505 bases: 46584080
Filtered reads: 2911505 bases: 46584080 (<18 bases long)
Analyzed reads: 0 bases: 0
::TOTAL::
Original reads: 2911505 bases: 46584080
Analyzed reads: 0 bases: 0
[samtools] faidx ./data/reference.fasta
REFERENCE: NC_008463
LENGTH: 6537648
libc++abi: terminating with uncaught exception of type std::domain_error: type must be number, but is null
Abort trap: 6
I have run the test drive successfully just before this, so I believe my install/setup is correct. Any guidance is appreciated!
Hi,
It looks like all of your reads are being filtered out, so breseq is trying to analyze zero reads.
Can you post the first few lines of your input FASTQ file PA14_dphz12_SRR11561537.1.fastq
so we can see why they are being rejected?
Thanks for the help. These data were downloaded from GenBank in their SRA archive format and I converted it to fastq using the default settings in their 'fastq-dump' tool in the SRA toolkit. Here are the first few lines of the file:
@SRR11561537.1.1 1 length=16
GCTCATGANGAGGATA
+SRR11561537.1.1 1 length=16
AAAAAEEE#AAAAEEE
@SRR11561537.1.2 2 length=16
GCTCATGANGAGGATA
+SRR11561537.1.2 2 length=16
AAAAAEEE#AAAAEEE
@SRR11561537.1.3 3 length=16
GCTCATGAAGAGGATA
+SRR11561537.1.3 3 length=16
AAAAAEEAAAAAAAEE
@SRR11561537.1.4 4 length=16
GCTCATGAAGAGGATA
+SRR11561537.1.4 4 length=16
AAAAAEE/AAAAAEEE
@SRR11561537.1.5 5 length=16
GCTCATGAAGAGGATA
+SRR11561537.1.5 5 length=16
AAAAAEEEAAAAAEEE
These reads are extremely short. breseq is throwing them all out because they are under it's normal minimum length requirement of 18 bases (as it says in the command line output).
You can make that cutoff smaller if you want, so they will be used, by adding --read-min-length 0
to your command. However, I don't think you'll see very good results in terms of calling mutations with such short reads. The minimum that we've ever really used breseq with is 36 bases.
OK, thanks for your help and insight!
Hello,
I'm new to breseq. I downloaded it from github, unzipped it and dragged and dropped it from my downloads folder to the folder I wanted it in (don't judge - I'm also new to bioinformatics and I had trouble navigating to my $PATH without being able to use miniconda). Within the breseq folder I created the test_drive folder and stored the 3 files: NC_012967.gbk, SRR030257_1.fastq, SRR030257_2.fastq. Note, I tried both the latest version of breseq (breseq-0.33.0-MacOSX-10.9+.tar.gz) and the most recent previous version (breseq-0.32.1-MacOSX-10.9.tar.gz).
I executed the test script using:
And this is the output I got:
I would appreciate any advice or pointers.
Thanks very much.