is our reads needs indexing before alignment like refrence genome or not ? some one explain to me plz

Question

[sabaghianamir70]

im new in this matter. and read some papers and watch some tutorial videos. i am in the way of aligning my dara. but in igv my data dosent match with my reference genome even iv got 92.7 % overal alignment with hisat2. some one help me pz

[Pierre Lindenbaum]

but in igv my data dosent match with my reference genome

what is "my data" ?

[sabaghianamir70]

Saccharomyces cerevisiae. i think its some thing wrong with my samtools

[cpad0112]

Probably you are loading wrong genome in IGV. Check reference version in IGV and it should match the reference you have used in alignment sabaghianamir70

[sabaghianamir70]

no i dont think so,. in igv i chose Saccharomyces cerevisiae refrence genome. both versions are same.

Answer 1

[h.mon]

Both reference genome in fasta format and mapped reads file in bam format need to be indexed, and the bam file needs to be sorted as well. For the fasta reference:

samtools faidx ref.fasta

For the mapped bam:

 samtools sort -o sorted.bam mapped.bam
 samtools index sorted.bam

In addition, the reference genome loaded into OGV need to be the same used for mapping.

[sabaghianamir70]

i think that is my problem , samtools wont work .im trying to install it for 2 days. its really annoying. i have the same problem as he had error installing samtools on ubuntu . but eventually is see this

libncurses5-dev : Depends: libtinfo5 (= 6.1-1ubuntu1) but 6.1-1ubuntu1.18.04 is to be installed Depends: libncurses5 (= 6.1-1ubuntu1) but 6.1-1ubuntu1.18.04 is to be installed Depends: libtinfo-dev (= 6.1-1ubuntu1) but it is not going to be installed

and the libtinfo-dev does not work.

[cpad0112]

If you are not able to install samtools, you can use igvtools (http://software.broadinstitute.org/software/igv/igvtools) for sorting and indexing. IGV ui has igvtools as menu option. If you have dependency issues in installation, try conda (user space installation). sabaghianamir70.