Saving unmapped reads from STAR

Question

[mihaell]

Hello,

I'm trying to save de unmapped reads from STAR aligner and I have some issues.

I would like to save the unmapped reads in a different output file. According to STAR Manual it's possible to save the unmapped reads using -outSAMunmapped Within parameter. Then the unmapped reads will be saved within Alignment.out.sam

I also tried the --outReadsUnmapped Fastx parameter and got Unmapped.out.mate1 and Unmapped.out.mate2. However I didn't understand how can I handle with these files.

I'm not sure about the results and I would like to know if exists another way to perform it.

Could anyone please let me know how to do this?

Thank you in advance

[Helen]

Hey Your question is exactly what I'm looking for I also tried the --outReadsUnmapped Fastx parameter and got Unmapped.out.mate1 , Unmapped.out.mate2.... until Unmapped.out.mate6 Can you tell me if it's right to use it? And why was it divided into 6 files?

Do you know any other way to extract the unmapped reads from STAR ?

Thanks in advance

[swbarnes2]

What's wrong with samtools view -hf 4 Aligned.sortedByCoord.out.bam > unmapped.bam. That's bam format, but you can change that to fastq easily enough.

[Helen]

thx, I tried what you said but I got an error : "open: No such file or directory [main_samview] fail to open "Aligned.sortedByCoord.out.bam" for reading."

P.S. my file is Alignment.out.sam

[swbarnes2]

Okay... what happens when you run that command line using the name of your file? And why are you keeping giant sam files around instead of bam files?

[Helen]

error: "[bam_header_read] EOF marker is absent. The input is probably truncated. [bam_header_read] invalid BAM binary header (this is not a BAM file). [main_samview] fail to read the header from "Aligned.out.sam"."

As for the sam file, I do not know STAR well and just wrote the commands I saw in the manual

Answer 1

[Devon Ryan]

Aren't the Unmapped.out.mate1 and Unmapped.out.mate2 files fastq files?