How to visualize a pair reads mapped genome.

Question

[K.Gee]

Hello! I'm looking to map some pair reads into a full viral genome. For visualizing the mapping genome, I am going to use IGV. I have a pair reads file generated from bowtie2 software, and I have the genome fasta file. I need someone to direct me or give an example of how to figure it out. I read the IGV manual, but it's a lit bit complicated to me.

[ATpoint]

You can load BAM files directly into the IGV. If you have a SAM file (the default output of bowtie2) use samtools view -o out.bam in.sam for conversion and then index the file with samtools index out.bam. Is this what you mean? If not please be more precise. Just a terminology thing, you have paired-end data, not pair reads ;-)

[K.Gee]

First of all, thanks for the prompt response. Actually yes! My main issue is that I cannot load my genome file into the IGV ( I forget mentioning that it a fasta file containing nucleotides). Some of IGVs manual links seem to be inactive. I will keep trying again, and I will bother you if Ill fail to load my files in the IGV if you still don't mind. Regarding the terminology, as I am a new kid on the block ( bioinformatics field), every correction, suggestion, and advice is acceptable :-).

[GenoMax]

I have a pair reads file generated from bowtie2 software

I hope that meant you have aligned data files because if you just have paired-end data (not aligned) then you will need to align it first in order to visualize it. You will need to use a user-defined genome and you will have to create it first. See this.

[K.Gee]

Thank you, too, for your response. Indeed, I did align the paired-end data. So I need to create the " user-defined genome." Unfortunately, some of IGV's links in the manual are inactive. I 'll make a try, and I hope I will make it.

[K.Gee]

I am gonna make a short recap: I have:

• a) a sam.file from bowtie2
• b) a bam file from this command: samtools view -o out.bam in.sam
• c) from samtools index out.bam -> i got this error [E::hts_idx_push] NO_COOR reads not in a single block at the end 0 -1 ( I will try to step over this error)
• d) I used the option genome from IGV tab menu (genome -> create.genome file -> I filled the genome's name, ID and I loaded the fast file of my genome ( containing nucleotides)

My question is how to load all together into a IGV software to get the visualized genome with the paired ends?

[GenoMax]

I will try to step over this error

No you can't do this with anything related to bioinformatics. You need to figure out the cause of error and fix it before your go forward. In this case the issue is you did not (co-ordinate) sort your BAM file before indexing it. So you should be doing

samtools sort -o sorted.bam in.sam
samtools index sorted.bam

If you correctely created the custom genome in IGV then at this point you just need to go "File --> Load From file --> Point to you sorted BAM". Remember to zoom in (may need to do it for a while) before you see aligned reads.

[K.Gee]

Actually that was the problem. I forgot to sort it first and I used different file names so I used a wrong file. I repeated the steps again so everything right now works fine for me. Just to mention on something. For the genome I used samtools faidx genome.fna before uploading to the IGV. Again thanks a lot for your answer and for your time.