samtools tview reference sequence

[rodney]

Does anyone else know how to get samtools tview to display the reference sequence at the top? I only see a string of N where I think the reference sequence should be.

[nilshomer]

Does anyone else know how to get samtools tview to display the reference sequence at the top? I only see a string of N where I think the reference sequence should be.

Add in the reference after the BAM file in the command line.

[Mansequencer]

Hi Rodney and Nilshomer,
I have a similar problem. I do add my reference fasta file after the BAM file but all I see are the Ns. I dont even see the reads that have aligned.
Any suggestions?
Thanks guys.

[nilshomer]

Hi Rodney and Nilshomer,
I have a similar problem. I do add my reference fasta file after the BAM file but all I see are the Ns. I dont even see the reads that have aligned.
Any suggestions?
Thanks guys.

Please give us what you typed into your command line.

[Mansequencer]

Here is the command:
samtools tview file.sort.bam reference.folded.fas

[nntao]

samtools tview issue

I have several issues with "samtools tview":
g : goto position, allows me to type a number, but not going anywhere
b : won't toggle

anyone has similar problem, and solutions

[nilshomer]

I have several issues with "samtools tview":
g : goto position, allows me to type a number, but not going anywhere
b : won't toggle

anyone has similar problem, and solutions

Enter the chromosome name and position (even if there is only one chromosome). For example "chr1:1000" (no quotes).

[naluru]

samtools tview

Hello,

I am having similar problem as mentioned here earlier. I have a string of N's in the reference genome, when I use the samtools tview command.Only the first 80 bases are present and remaining are all NNNNNN's. I tried using g option and tried different chromosome numbers. Still I have only N's.

Any help will be highly appreciated.

Here is the command I gave:

samtools tview MH_0001alignedreadssorted.bam danRer6.fa

[epigen]

If your reference sequence is correct and indexed, as well as you BAM file, you should see the reference sequence where reads aligned. I noted that if there are no reads for a long stretch of bases, tview doesn't bother to show the reference, it shows only Ns. When I checked with another BAM file that had reads at that location, everything was OK. Before I thought the fasta file was currupted - which might very well be a reason for only seeing Ns.

[tonge]

Retracted message

[qtrinh]

Did you use 'danRer6.fa' in your alignment step? I think the issue here is that the reference used in the alignment step is not the same as 'danRer6.fa'

Hello,

I am having similar problem as mentioned here earlier. I have a string of N's in the reference genome, when I use the samtools tview command.Only the first 80 bases are present and remaining are all NNNNNN's. I tried using g option and tried different chromosome numbers. Still I have only N's.

Any help will be highly appreciated.

Here is the command I gave:

samtools tview MH_0001alignedreadssorted.bam danRer6.fa

[swbarnes2]

It's probably not a problem with tview, you can get the same issue in mpileup. Double-check that the names in your reference file are exactly the same as in your .sam file, or that there aren't any odd characters that might be confusing things.

[VeBeKay]

For me, it was a colon ":" in my chromosome name for the reference sequence. When I deleted the colon, the actual sequence showed up (before that it was only N's).

[Tally]

Thank you VeBeKay!! That solved my problem perfectly :-) However I had to delete it in the reference AND rerun the alignment and subsequent steps, time-consuming but 100% effective.
I am surprised this was an issue, as the fasta reference was created using samtools faidx region command, which utilises a semi-colon to define the region. I tried faidx region extract with no semi-colon, and this does not give the correct output. Does anyone know of how to use faidx to extract regions to a multi-fasta which does not have semi-colons in the output files? It seems silly to require them for faidx yet malfunction over them in tview.

[HeidiJTP]

I am also having this problem, where the first 80 bases are present, then mostly Ns except in a few positions where aligned reads appear. I am just running a test set to try to learn the process, so I took a reference sequence and fragmented it into 500 bp segments with 50 bp overlaps, then aligned the fragments (short_reads.fas) back to the reference.fas. Thus, the full reference should have fragments aligned to it. Here are the commands I used:

$ bwa index reference.fas
$ bwa aln reference.fas short_reads.fas >short_reads.sai
$ bwa samse reference.fas short_reads.sai short_reads.fas >short_reads.sam
$ samtools faidx reference.fas
$ samtools import reference.fas.fai short_reads.sam short_reads.bam
$ samtools sort short_reads.bam short_reads.srt
$ samtools index short_reads.srt.bam
$ samtools tview short_reads.srt.bam reference.fas

I made sure that there were no colons in any sequence titles, but I am not sure I have used all the commands correctly. I would really appreciate any help!