Actually, I used the bam2fastq tool, but as I said, I also got "!" score for each base.
My data are pretty new.
Hello,
I got reads from PacBio sequencing. The reads are in BAM format. I converted them into FASTQ format.
I both used the bam2fastq and samtools fastq tools to do that.
The problem I have is that I got the " ! " score for all bases of the sequences with both tools, which means the bases are all wrong. What is not good because the phred score I got with FASTQC is really good (and reads obtained with the PacBio tech are usually always good)
Any idea?
Bests
I got reads from PacBio sequencing. The reads are in BAM format.
Then use PacBio's utility bam2fastx to do the conversion.
The problem I have is that I got the " ! " score for all bases of the sequences with both tools, which means the bases are all wrong.
There is a thread on SeqAnswers about your observation. I will deep link one post. You can read the entire thread. Is your data older?
Actually, I used the bam2fastq tool, but as I said, I also got "!" score for each base.
My data are pretty new.
I guess the ! has not been replaced with meaningful values as stated by user rhall (who works for PacBio) in another post in the thread I had linked above from SeqAnswers.
I would follow @lieven's advice above or replace ! with something else using reformat.sh from BBMap suite.
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base scores in pacbio fastq files have no to very little meaning due to the specifics of the pacbio technique (at least that's how I remember from older datasets, perhaps it changed for more recent datasets), so don't worry too much about this I would say.
If you use the conversion tools of pacbio smrt package, I think you can even say what you want to scores to be
Just use the data as it is without taking the scores into account.
some more info which I recently found online: