how to split forward and reverse reads from sam file

Question

[Bioinfo]

Hello everyone,

I have question please

I have bam file containing unmapped reads that I want to assemble.

I tried samtools fastq to obtain the two files forward and reverse

I did this command

samtools fastq -1 wPru_1_Unmapped_reads.fastq -2 wPru_2_Unmapped_reads.fastq   wPru_Unmapped_reads.bam

but when I wanted to perform the assembly (shovill) it didn't work it shows that the number of forward is larger than the number of reverse reads

The number of left read-pairs is larger than the number of right read-pairs

my objective now is how to extract forward and reverse reads from bam file

Thank you very much

[lieven.sterck]

How did you obtain the bam file with unmapped reads?

What I suspect is that in the unaligned bam file there will be reads pairs for which one is mapped and the other not , and if the bam was created not taking this into account it might not work the split them in equal numbers.

[Bioinfo]

Thank you very much for your answer i did this command to obtain unmapped reads

samtools view -h -f 4 nom_de_souche_Resultat.bam > nom_de_souche_Unmapped_reads.bam

i save all the files so i can fix the error in any step i mapped my reads against the eference using bowtie2 after that i ransfer the sam file to bam file and i extracted the unmapped reads from bam file then ai tranformed bam file to two fastq files using the command in the question

[lieven.sterck]

don't know the specifics from the top of my head but I don't think that the samtools cmdline will do this correctly. If will process all reads as separate entities (so not taking the PE info into account).

alternatively have a look at the bbmap software, that you can specificly configure to write out all kinds of read mapping results

[Bioinfo]

Okay , thank you very much

[lieven.sterck]

A small educational note: I added (code) markup to your post for increased readability. You can do this by selecting the text and clicking the 101010 button. When you compose or edit a post that button is in your toolbar, see image below:

Answer 1

[GenoMax]

As noted by @lieven you could use reformat.sh from BBMap suite..

reformat.sh in=your.sam out1=R1.fq out2=R2.fq

Consider following additional options to add :

mappedonly=f            Toss unmapped reads.
unmappedonly=f          Toss mapped reads.
pairedonly=f            Toss reads that are not mapped as proper pairs.
unpairedonly=f          Toss reads that are mapped as proper pairs.
primaryonly=f           Toss secondary alignments.  Set this to true for sam to fastq conversion.