starWithAnn in 1.4.9

[alexyfyf]

RNAsik version 1.4.9

fyan0011@biomate1:~/project/GFPhvsl/data/20180416Klf$ RNAsik -fqDir ../Klf_HetvsNull_rnaseq/ -align starWithAnn -counts -metrics -threads 10 -fastaRef ~/ref-seq/ensembl/mus/Mus_musculus.GRCm38.dna.primary_assembly.fa -extn .fastq.gz -gtfFile ~/ref-seq/ensembl/mus/Mus_musculus.GRCm38.91.gtf
00:00:00.034    Error (/mnt/software/apps/RNAsik-pipe-1.4.9/bin/../src/sikSanityCheck.bds, line 87, pos 73): Usage: -gtfFile /path/to/GTFfile

fyan0011@biomate1:~/project/GFPhvsl/data/20180416Klf$ RNAsik -fqDir ../Klf_HetvsNull_rnaseq/ -align starWithAnn -counts -metrics -threads 10 -refFiles ../20180109ensembl/sikRun/refFiles/ 
00:00:00.046    Error (/mnt/software/apps/RNAsik-pipe-1.4.9/bin/../src/sikSanityCheck.bds, line 87, pos 73): Usage: -gtfFile /path/to/GTFfile

Hi, I found RNAsik 1.4.9 cannot recognize the gtf files for starwithann aligner. The same thing happens with 1.5.0. Can you please have a look?

Cheers,

Alex

[alexyfyf]

RNAsik version 1.5.0

fyan0011@biomate1:~/project/GFPhvsl/data/20180416Klf$ ~/software/RNAsik-pipe/bin/RNAsik -fqDir ../Klf_HetvsNull_rnaseq/ -align starWithAnn -gtfFile ../20180109ensembl/sikRun/refFiles/Mus_musculus.GRCm38.91.gtf
00:00:00.038    Error (/persistent/home/fyan0011/software/RNAsik-pipe/bin/../src/sikSanityCheck.bds, line 45, pos 32): You need to either specify -genomeIdx or -fastaRef or -refFiles

fyan0011@biomate1:~/project/GFPhvsl/data/20180416Klf$ ~/software/RNAsik-pipe/bin/RNAsik -fqDir ../Klf_HetvsNull_rnaseq/ -align starWithAnn -refFiles ../20180109ensembl/sikRun/refFiles/  
00:00:00.061    Error (/persistent/home/fyan0011/software/RNAsik-pipe/bin/../src/sikSanityCheck.bds, line 50, pos 9): Usage: -refFiles /path/to/refFiles/directory OR -gtfFile /path/to/GTFfile

Even with the update in https://github.com/alexyfyf/RNAsik-pipe/commit/f322e58a3ff59e4e84030a890a3552bd9339a7d0, still having this issue. slightly different.

[serine]

Thanks for reporting this.

So you seem to have two different errors there. Let's focus on the first error you've reported and stick to version 1.5.0.

So I think OR (||) is also wrong here, because even even if you pass -refFile it skips to the second conditional and fails on that because of the OR. We need to re-wire the logic.

Basically what I'd like is to either use -refFiles OR -gtfFile And that sanity check is to ensure that either of those two options are used if user wants to use starWithAnn

If you could try to re-write some of the logic to meet that criteria and send me PR that would be much appreciated.

Thanks

p.s I've edited your comments to display error message, could you use that style of code reporting, makes it easier to read and follow, cheers

[alexyfyf]

Successfully fool the sanitycheck, but STAR still didn't take the gtf fles. https://github.com/alexyfyf/RNAsik-pipe/commit/6861ffebdc8a894da302471ba1960505850106e9

~/software/RNAsik-pipe/bin/RNAsik -fqDir ../Klf_HetvsNull_rnaseq/ -align starWithAnn -gtfFile ../20180109ensembl/sikRun/refFiles/Mus_musculus.GRCm38.91.gtf -fastaRef ../20180109ensembl/sikRun/refFiles/Mus_musculus.GRCm38.dna.primary_assembly.fa

The script from the html file

Task hint          : 'STAR --runThreadN 10; --runMode alignReads; --genomeDir sikRun/refFiles/Mus_musculus.GRCm38.dna.primary_assembly.starIdx; --outSAMattrRGline ID:001 CN'

No gtf files taken, although it passed the sanity check.

[serine]

Task hint wouldn't show full command ran. Can paste what command from BAM's file header? BAMs header will show exact parameters used to generate that BAM file.

Also can you please check sikRun/logs/toolsOpts/ and see if there is an entry for STAR

[serine]

Also if you want me to pull your fix in, you should fix it properly, but if you are not aiming for PR then any hack will do.

I'd appreciate a PR as this needs fixing, I just don't have time to address this issue now

Cheers

[alexyfyf]

fyan0011@biomate1:~/project/GFPhvsl/data/20180416Klf/sikRun/bamFiles$ ~/software/RNAsik-pipe/bin/RNAsik -fqDir ../Klf_HetvsNull_rnaseq/ -align starWithAnn -gtfFile ../20180109ensembl/sikRun/refFiles/Mus_musculus.GRCm38.91.gtf -fastaRef ../20180109ensembl/sikRun/refFiles/Mus_musculus.GRCm38.dna.primary_assembly.fa -genomeIdx ../20180109ensembl/sikRun/refFiles/Mus_musculus.GRCm38.dna.primary_assembly.starIdx/

The command I use, with fixed script that fools the sanitycheck. The results bam files header @PG line.

@PG     ID:STAR PN:STAR VN:STAR_2.5.2b  CL:STAR   --runMode alignReads   --runThreadN 10   --genomeDir ../20180109ensembl/sikRun/refFiles/Mus_musculus.GRCm38.dna.primary_assembly.starIdx/   --readFilesIn /persistent/home/fyan0011/project/GFPhvsl/data/20180416Klf/../Klf_HetvsNull_rnaseq/Klf1_Het_Rep1_R1.fastq.gz      --readFilesCommand zcat      --outFileNamePrefix sikRun/alignerFiles/Klf1_Het_Rep1_   --outSAMtype BAM   Unsorted      --outSAMunmapped Within      --outSAMattrRGline ID:001   CN:Unknown   DS:RNA-seq   PL:ILLUMINA   PM:MiSeq   SM:Klf1_Het_Rep1