problem for setting BWA arguments for alignment

Question

[modarzi]

Hi,

I want to align my samples by BWA. my samples are RNA-seq and Single-End. as I read on Manual Reference Pages - bwa (1), I have to run below command:

bwa mem [-aCHMpP] [-t nThreads] [-k minSeedLen] [-w bandWidth] [-d zDropoff] [-r seedSplitRatio] [-c maxOcc] [-A matchScore] [-B mmPenalty] [-O gapOpenPen] [-E gapExtPen] [-L clipPen] [-U unpairPen] [-R RGline] [-v verboseLevel] db.prefix reads.fq [mates.fq]

all arguments in [ ] are optional but my question is about "db.prefix" argument.I don't know what it is and how can provide it? based on alignment logic, it should be reference genome but I am not sure and if it is reference I don't know how to prepare it? I appreciate if you share your comment with me.

Best Regards,

Mohammad

[WouterDeCoster]

bwa is not an appropriate aligner for RNA-seq (unless it's prokaryotic), you should use a splice-aware aligner such as STAR or HISAT2.

Answer 1

[Tm]

For using bwa, you need two things:

1. reads file
2. reference sequence/s on which you want to map the reads

Now, first before using 'mem' for mapping, you have to index the reference sequences using 'bwa index' followed by bwa mem where you will have to provide a prefix of reference file indexed. For example:

bwa index ref.fa (it will generate ref.fa.amb, ref.fa.ann, ref.fa.bwt, ref.fa.pac, ref.fa.sa files )
bwa mem ref.fa reads_SE.fastq -t 30 > reads_on_reference.sam

So, here in above command "ref.fa" is your db.prefix.

[modarzi]

Thanks.

my data set belong to human. I consruct bwa index(wg.fa) and for alignment I run below command:

bwa mem ~/cirRNA/apps/wg.fa sra_data_SRR1427482.fastq -t 30 > sra_data_SRR1427482_bwa.sam

and then in my terminal I have below result:

[M::bwa_idx_load_from_disk] read 0 ALT contigs

[M::process] read 8646532 sequences (300000008 bp)...

but after some minutes, my process will killed.

I also, ran this command for another sample and receive below message:

[M::bwa_idx_load_from_disk] read 0 ALT contigs

Segmentation fault (core dumped)

I appreciate if you share your comment with me.

Best Regards,

Mohammad