Error with RNA-SeQC

[angerusso]

Hi,

I used following 4 steps to get sequencing statistics from RNASeqC but I get error, could anyone help me understand and get rid of this error?

Thanks.

RNA-SeQC v1.1.7 05/14/12
Creating rRNA Interval List based on given GTF annotations
java.lang.ArrayIndexOutOfBoundsException: 1
at org.broadinstitute.cga.rnaseq.RNASeqMetrics$MetricSample.readInSamples(RNASeqMetrics.java:1369)
at org.broadinstitute.cga.rnaseq.RNASeqMetrics.prepareFiles(RNASeqMetrics.java:182)
at org.broadinstitute.cga.rnaseq.RNASeqMetrics.execute(RNASeqMetrics.java:165)
at org.broadinstitute.cga.rnaseq.RNASeqMetrics.main(RNASeqMetrics.java:135)
RNA-SeQC Total Runtime: 0 min
-----------------

java -jar picard-tools/v1.87/AddOrReplaceReadGroups.jar I=MCF7/accepted_hits.bam O=MCF7/accepted_hits_readgrps.bam VALIDATION_STRINGENCY=LENIENT SORT_ORDER=coordinate RGID=MCF7 RGCN=MCF7 RGLB=lane6 RGPL=illumina RGPU=lane6 RGSM=accepted_hits.bam

java -jar picard-tools/v1.87/SortSam.jar I=MCF7/accepted_hits_readgrps.bam O=MCF7/accepted_hits_readgrps_sorted.bam SO=coordinate

java -jar picard-tools/v1.87/ReorderSam.jar I=MCF7/accepted_hits_readgrps_sorted.bam O=MCF7/accepted_hits_readgrps_sorted.reordered.bam R=refGenome/hg19.fa CREATE_INDEX=true

java -jar softwares/RNA-SeQC_v1.1.7.jar -n 1000 -s MCF7/accepted_hits_readgrps_sorted.reordered.bam -t index/Homo_sapiens/UCSC/hg19/Annotation/Archives/archive-2012-03-09-03-24-41/Genes/genes.gtf -r refGenome/hg19.fa -o reportMCF7

[pkachroo]

I get exactly the same error

RNA-SeQC v1.1.7 05/14/12
Creating rRNA Interval List based on given GTF annotations
java.lang.ArrayIndexOutOfBoundsException: 1
at org.broadinstitute.cga.rnaseq.RNASeqMetrics$MetricSample.readInSamples(RNASeqMetrics.java:1369)
at org.broadinstitute.cga.rnaseq.RNASeqMetrics.prepareFiles(RNASeqMetrics.java:182)
at org.broadinstitute.cga.rnaseq.RNASeqMetrics.execute(RNASeqMetrics.java:165)
at org.broadinstitute.cga.rnaseq.RNASeqMetrics.main(RNASeqMetrics.java:135)
RNA-SeQC Total Runtime: 1 min

I do not understand the reason for this error, I also noticed that the header of my original bam file had 3 columns and after i ran reordersam, the columns in bam file increased, could this be a problem. I am just pasting few lines:
ORIGINAL FILE:
@HD VN:1.0 SO:coordinate
@SQ SN:chr1 LN:195471971
@SQ SN:chr10 LN:130694993
@SQ SN:chr11 LN:122082543
@SQ SN:chr12 LN:120129022
@SQ SN:chr13 LN:120421639
@SQ SN:chr14 LN:124902244
@SQ SN:chr15 LN:104043685
@SQ SN:chr16 LN:98207768
@SQ SN:chr17 LN:94987271
@SQ SN:chr18 LN:90702639
@SQ SN:chr19 LN:61431566
@SQ SN:chr1_GL456210_random LN:169725
@SQ SN:chr1_GL456211_random LN:241735
@SQ SN:chr1_GL456212_random LN:153618
@SQ SN:chr1_GL456213_random LN:39340

AFTER REORDERING:
@HD VN:1.0 SO:coordinate
@SQ SN:chr1 LN:195471971 UR:file:/work_Fid/sukmb277/references/mouse_mm10/mm10.fa M5:c4ec915e7348d42648eefc1534b71c99
@SQ SN:chr10 LN:130694993 UR:file:/work_Fid/sukmb277/references/mouse_mm10/mm10.fa M5:7831ecda5dd6bcf838e2452ea0139eac
@SQ SN:chr11 LN:122082543 UR:file:/work_Fid/sukmb277/references/mouse_mm10/mm10.fa M5:e168c7a3194813f597181f26bb1bd13f
@SQ SN:chr12 LN:120129022 UR:file:/work_Fid/sukmb277/references/mouse_mm10/mm10.fa M5:671f85bb54a6e097d631e2e2dd813ad4
@SQ SN:chr13 LN:120421639 UR:file:/work_Fid/sukmb277/references/mouse_mm10/mm10.fa M5:7f9b9fa3fbd9a38634107dfdc7fd8dc8
@SQ SN:chr14 LN:124902244 UR:file:/work_Fid/sukmb277/references/mouse_mm10/mm10.fa M5:bf4e1efa25a8fd23b41c91f9bcb86388
@SQ SN:chr15 LN:104043685 UR:file:/work_Fid/sukmb277/references/mouse_mm10/mm10.fa M5:106358dace00e5825ae337c1f1821b5e
@SQ SN:chr16 LN:98207768 UR:file:/work_Fid/sukmb277/references/mouse_mm10/mm10.fa M5:5482110a6cedd3558272325eee4c5a17
@SQ SN:chr17 LN:94987271 UR:file:/work_Fid/sukmb277/references/mouse_mm10/mm10.fa M5:0d21e8edbfcd8410523b2b94e6dae892
@SQ SN:chr18 LN:90702639 UR:file:/work_Fid/sukmb277/references/mouse_mm10/mm10.fa M5:46fda2f7e6dbf91bff91d6703e004afb
@SQ SN:chr19 LN:61431566 UR:file:/work_Fid/sukmb277/references/mouse_mm10/mm10.fa M5:7d363594531514ce41dfacfd97bc995d
@SQ SN:chr1_GL456210_random LN:169725 UR:file:/work_Fid/sukmb277/references/mouse_mm10/mm10.fa M5:0cc560d98f9f22f4385397db82e1c108
@SQ SN:chr1_GL456211_random LN:241735 UR:file:/work_Fid/sukmb277/references/mouse_mm10/mm10.fa M5:36e85680c669756c9a1554cf31c9de03
@SQ SN:chr1_GL456212_random LN:153618 UR:file:/work_Fid/sukmb277/references/mouse_mm10/mm10.fa M5:4c4dc3bfe987e3bc4ef4756bef269373
@SQ SN:chr1_GL456213_random LN:39340 UR:file:/work_Fid/sukmb277/references/mouse_mm10/mm10.fa M5:d4cb9051fe171205dd39980e110bf63e
@SQ SN:chr1_GL456221_random LN:206961 UR:file:/work_Fid/sukmb277/references/mouse_mm10/mm10.fa M5:e21da65d7276b256b8edf92660a928b0

But i did follow the same steps as mentioned on :

https://sites.google.com/site/riverlee2008/post/tosuccessfullyrunrna-seqcontophatalignmentresult

and then ran RNA-SeQC v1.1.7 , My Java version on cluster is: java version "1.6.0_31"
Java(TM) SE Runtime Environment (build 1.6.0_31-b04)
Java HotSpot(TM) 64-Bit Server VM (build 20.6-b01, mixed mode)

Could anyone help me to know the cause of this problem ?

Thanks a lot,

regards,

Priya

[rnastar]

I am also having the same problem, except with a bam file created with STAR output. Anyone have any ideas?

[Timothy Amos]

The error message says there is a problem with "readInSamples"

The correct use of the -s option is, according to http://www.broadinstitute.org/cancer/cga/rnaseqc_run:

-s <arg>

Sample File: tab-delimited description of samples and their bams. This file header is:
Sample ID Bam File Notes
When running on just one sample, this argument can be a string of the form
"Sample ID|Bam File|Notes", where Bam File is the path to the input file.

Rather than

-s MCF7/accepted_hits_readgrps_sorted.reordered.bam

I got this error by accidentally having '\t' in my tab-delimited samples file rather than a literal tab. My mistake was to use:

echo "Sample ID\tBam File\tNotes" > ${OUTDIR}/Samples.txt

rather than including the -e option:

echo -e "Sample ID\tBam File\tNotes" > ${OUTDIR}/Samples.txt