STAR 2.4.0k Missing SA and index file in GenomeForPass2

[Carina Jønck]

Hello.

I am running STAR pass2 according to the script: https://code.google.com/p/rna-star/issues/attachmentText?id=7&aid=70001000&name=STAR_2pass.sh&token=ABZ6GAcYRkqv2DT118nT59vVuV0ihUzPvw%3A1427729052975

After having some trouble with memory (see other question) we downloaded the new version 2.4.0k. This is very fast (seconds instead of hours?). However, it does not give the output needed for further analysis as the SA and index file in GenomeForPass2 are missing. See I/O below.

How do I get these files?

Cheers
Carina

 

[gsp720@ri-ngs-centos02 Pass1]$ /mnt/NGS01/biss/bin/STARstatic --runThreadN 1 --outSAMattributes All --genomeLoad LoadAndKeep --genomeDir /mnt/NGS01/biss/tasks/task_45/hg19/ --readFilesIn ../trimmed.fastq

Apr 08 10:54:08 ..... Started STAR run

Apr 08 10:54:08 ..... Started mapping

Apr 08 10:54:21 ..... Finished successfully

[gsp720@ri-ngs-centos02 Pass1]$ cd ..

[gsp720@ri-ngs-centos02 pool1_004]$ cd GenomeForPass2/

[gsp720@ri-ngs-centos02 GenomeForPass2]$ /mnt/NGS01/biss/bin/STARstatic --runThreadN 1 --outSAMattributes All --genomeLoad LoadAndKeep --genomeDir /mnt/NGS01/biss/tasks/task_45/hg19/ --readFilesIn ../trimmed.fastq

Apr 08 10:55:39 ..... Started STAR run

Apr 08 10:55:39 ..... Started mapping

Apr 08 10:55:53 ..... Finished successfully

[gsp720@ri-ngs-centos02 GenomeForPass2]$ cd ..

[gsp720@ri-ngs-centos02 pool1_004]$ cd Pass2

[gsp720@ri-ngs-centos02 Pass2]$ /mnt/NGS01/biss/bin/STARstatic --runThreadN 1 --outSAMattributes All --genomeLoad LoadAndKeep --outFilterMultimapNmax 1 --genomeDir ../GenomeForPass2 --readFilesIn ../trimmed.fastq

Apr 08 10:58:03 ..... Started STAR run

 

EXITING because of FATAL ERROR: could not open genome file ../GenomeForPass2/SA

 

SOLUTION: check that the path to genome files, specified in --genomDir is correct and the files are present, and have user read permsissions

 

Apr 08 10:58:03 ...... FATAL ERROR, exiting

[gsp720@ri-ngs-centos02 Pass2]$ cd ..

[gsp720@ri-ngs-centos02 pool1_004]$ cd GenomeForPass2/

[gsp720@ri-ngs-centos02 GenomeForPass2]$ ls

Aligned.out.sam    Genome                sjdbInfo.txt

chrLength.txt      genomeParameters.txt  sjdbList.out.tab

chrNameLength.txt  Log.final.out         SJ.out.tab

chrName.txt        Log.out               SJ.out.tab.Pass1.sjdb

chrStart.txt       Log.progress.out

[Alexander Dobin]

Hi Carina,

this 2-pass script is outdated and will not work with the current version of STAR.

Moreover, if you do not use annotations, you can run an "automated" 2-pass simply using --twopass1readsN -1, e.g.

STAR --runThreadN 1 --outSAMattributes All --genomeDir /mnt/NGS01/biss/tasks/task_45/hg19/ --readFilesIn ../trimmed.fastq --sjdbOverhang 100 --twopass1readsN -1

What was the command you used for generating /mnt/NGS01/biss/tasks/task_45/hg19/ genome?

It also seems that your 1st pass mapping did not complete, since it only took a few seconds.

How did you trim your reads? If trimming left empty sequences, it would terminate mapping.

Please try to run simple 1-pass STAR command on untrimmed reads first.

Cheers

Alex

[Carina Jønck]

Hi Alex

 

Thank you for your quick reply :)

 

I still have problems when I use your suggestions, but now it is with memory, so I put it on an earlier post.

Cheers

Carina