RNA-SeQC IntronicExpressionReadBlock error

Question

[konradgru]

Hi all,

I have got an error when running RNA-SeQC I made sure to have all prerequesites: coordinate sorted bam, bam is indexed (bai), bam are duplicate marked (picard tools), bam has read groups (picard tools), bam chr are ordered in reference order (picard ReorderSam), reference indexed (faid), reference dictionary (dict)

I made my own rRNA list file:

cat Homo_sapiens.GRCh38.84.gtf|grep 'gene_biotype "rRNA"' |awk '$3 == "transcript"' |cut -f1,4,5,7,9 > rRNA.list

and added the sam header.

RNA-SeQC can only handle gtf lines with transcript_id in field 9; thus:

cat Homo_sapiens.GRCh38.84.gtf |grep transcript_id > Homo_sapiens.GRCh38.84-transcript_idLines.gtf

... yes, RNA-SeQC makes us jump a lot of hoops.

Still, when running this line:

java -jar RNA-SeQC_v1.1.8.jar -o RNASeqQC -r HS_GRCH38_84_ucsc-all.fa -s "28129|accepted_hits_coordSorted_1Primary_mk_dup_RG_chrReordered.bam|no notes" -t Homo_sapiens.GRCh38.84-transcript_idLines.gtf -rRNA rRNA-chr.list

I get lots of cryptic exceptions during "IntronicExpressionReadBlock Walker":

RNA-SeQC v1.1.8.1 07/11/14
Retriving contig names from reference
         contig names in reference: 455
Loading GTF for Read Counting
Converting to refGene
Transcript objects to RefGen format:    4 s
Running IntronicExpressionReadBlock Walker ....
Exception in thread "main" java.lang.ExceptionInInitializerError
        at org.broadinstitute.sting.gatk.GenomeAnalysisEngine.<init>(GenomeAnalysisEngine.java:146)
        at org.broadinstitute.sting.gatk.CommandLineExecutable.<init>(CommandLineExecutable.java:53)
        at org.broadinstitute.sting.gatk.CommandLineGATK.<init>(CommandLineGATK.java:55)
        at org.broadinstitute.cga.rnaseq.gatk.GATKTools.runIntronReadCount(GATKTools.java:216)
        at org.broadinstitute.cga.rnaseq.ReadCountMetrics.runRegionCounting(ReadCountMetrics.java:244)
        at org.broadinstitute.cga.rnaseq.ReadCountMetrics.runReadCountMetrics(ReadCountMetrics.java:59)
        at org.broadinstitute.cga.rnaseq.RNASeqMetrics.runMetrics(RNASeqMetrics.java:225)
        at org.broadinstitute.cga.rnaseq.RNASeqMetrics.execute(RNASeqMetrics.java:171)
        at org.broadinstitute.cga.rnaseq.RNASeqMetrics.main(RNASeqMetrics.java:139)
Caused by: java.lang.RuntimeException: java.util.concurrent.ExecutionException: java.lang.RuntimeException: could    not create class file from NativeRegExp$Prototype.class
        at org.reflections.Reflections.scan(Reflections.java:166)
        at org.reflections.Reflections.<init>(Reflections.java:91)
        at org.broadinstitute.sting.utils.classloader.PluginManager.<clinit>(PluginManager.java:79)
        ... 9 more
Caused by: java.util.concurrent.ExecutionException: java.lang.RuntimeException: could not create class file from  NativeRegExp$Prototype.class
        at java.util.concurrent.FutureTask.report(FutureTask.java:122)
        at java.util.concurrent.FutureTask.get(FutureTask.java:192)
        at org.reflections.Reflections.scan(Reflections.java:162)
        ... 11 more
Caused by: java.lang.RuntimeException: could not create class file from NativeRegExp$Prototype.class
        at org.reflections.scanners.AbstractScanner.scan(AbstractScanner.java:41)
        at org.reflections.Reflections$2.run(Reflections.java:149)
        at java.util.concurrent.Executors$RunnableAdapter.call(Executors.java:511)
        at java.util.concurrent.FutureTask.run(FutureTask.java:266)
        at java.util.concurrent.ThreadPoolExecutor.runWorker(ThreadPoolExecutor.java:1142)
        at java.util.concurrent.ThreadPoolExecutor$Worker.run(ThreadPoolExecutor.java:617)
        at java.lang.Thread.run(Thread.java:745)
Caused by: java.io.IOException: invalid constant type: 15
        at javassist.bytecode.ConstPool.readOne(ConstPool.java:1023)
        at javassist.bytecode.ConstPool.read(ConstPool.java:966)
        at javassist.bytecode.ConstPool.<init>(ConstPool.java:127)
        at javassist.bytecode.ClassFile.read(ClassFile.java:693)
        at javassist.bytecode.ClassFile.<init>(ClassFile.java:85)
        at org.reflections.adapters.JavassistAdapter.createClassObject(JavassistAdapter.java:86)
        at org.reflections.adapters.JavassistAdapter.createClassObject(JavassistAdapter.java:22)
        at org.reflections.scanners.AbstractScanner.scan(AbstractScanner.java:38)
        ... 6 more

Has anyone experienced such a problem? Or can anyone advice me how to track down the cause of this error?

Thanks for any help

Answer 1

[iarmean]

Had the same error within similar context. The fix for mine was to use Java-1.7, was using Java-1.8 as default. Maybe this fixes yours too!