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CollectRnaSeqMetrics shows "Sequence dictionaries differ" error

Hi all! I was trying to run the CollectRnaSeqMetrics from Picard tools. My command was:

java -Xmx4g -jar picard.jar CollectRnaSeqMetrics \
I=/home/ania/STAR_results/SLX-10113.D701_D505.C80D1ANXX/Aligned.out.sam \
O=/home/ania/mappingQC/1-5.RNA_Metrics \
REF_FLAT=/home/ania/STAR/STAR-2.5.2a/bin/Linux_x86_64/gtf/refFlat.txt \
REFERENCE_SEQUENCE=/home/ania/STAR/STAR-2.5.2a/bin/Linux_x86_64/genome/GRCh38_r85.all.fa \
STRAND_SPECIFICITY=NONE \
RIBOSOMAL_INTERVALS=/home/ania/mappingQC/gencode.v19.rRNA.interval_list \
CHART_OUTPUT=/home/ania/mappingQC/file.rnaseq.pdf

I created Ribosomal_intervals file by using this script: script and making a few changes in it, namely: I changed hg19 to hg38 and gene_type to transcript_biotype

For mapping I used the STAR tool. Unfortunatelly, what I get is an error:

Exception in thread "main" picard.PicardException: Sequence dictionaries differ in /home/ania/STAR_results/SLX-10113.D701_D505.C80D1ANXX/Aligned.out.sam and /home/ania/mappingQC/gencode.v19.rRNA.interval_list

Together with many warnings like these:

WARNING 2016-07-27 11:06:36 IntervalList Ignoring interval for unknown reference: 13:60613608-60613734 + ENST00000411144

WARNING 2016-07-27 11:06:36 IntervalList Ignoring interval for unknown reference: Y:18348035-18348147 - ENST00000516858

Such warnings refere to all chromosomes.

I don't know what to do with this problem. Could you please tell me how I can solve this? I would be extremely grateful.

collectrnaseqmetrics picard
• 4,061 views • link
updated by GenoMax GenoMax • written by anjasta47 • anjasta47

At least for the Sequence dictionaries differ part see the solution in this thread. It appears that you used the same script for creating intervals file so it should be applicable in your case.

• 1 views • link
written by GenoMax GenoMax

I have seen this thread while looking for the solution but it doesn't solve my problem because both my sam file and intervals file are tab-delimited.

• 1 views • link
written by anjasta47 • anjasta47

1 answer

show me the BAM header for samtools view -H Aligned.out.sam and the header for gencode.v19.rRNA.interval_list

• 1 views • link
written by Pierre Lindenbaum Pierre Lindenbaum 
@HD     VN:1.4
@SQ     SN:10   LN:133797422
@SQ     SN:11   LN:135086622
@SQ     SN:12   LN:133275309
@SQ     SN:13   LN:114364328
@SQ     SN:14   LN:107043718
@SQ     SN:15   LN:101991189
@SQ     SN:16   LN:90338345
@SQ     SN:17   LN:83257441
@SQ     SN:18   LN:80373285
@SQ     SN:19   LN:58617616
@SQ     SN:1    LN:248956422
@SQ     SN:20   LN:64444167
@SQ     SN:21   LN:46709983
@SQ     SN:22   LN:50818468
@SQ     SN:2    LN:242193529
@SQ     SN:3    LN:198295559
@SQ     SN:4    LN:190214555
@SQ     SN:5    LN:181538259
@SQ     SN:6    LN:170805979
@SQ     SN:7    LN:159345973
@SQ     SN:8    LN:145138636
@SQ     SN:9    LN:138394717
@SQ     SN:MT   LN:16569
@SQ     SN:X    LN:156040895
@SQ     SN:Y    LN:57227415
@PG     ID:STAR PN:STAR VN:STAR_2.5.2a  CL:/home/ania/STAR/STAR-2.5.2a/bin/Linux_x86_64/STAR   --runThreadN 36   --genomeDir /home/ania/STAR/STAR-2.5.2a/bin/Linux_x86_64/genome_index   --readFilesIn /home/ania/data_FastQ/SLX-10113.D701_D505.C80D1ANXX.fq
@CO     user command line: /home/ania/STAR/STAR-2.5.2a/bin/Linux_x86_64/STAR --runThreadN 36 --genomeDir /home/ania/STAR/STAR-2.5.2a/bin/Linux_x86_64/genome_index --readFilesIn /home/ania/data_FastQ/SLX-10113.D701_D505.C80D1ANXX.fq

@SQ     SN:chr1 LN:248956422    AS:hg38
@SQ     SN:chr2 LN:242193529    AS:hg38
@SQ     SN:chr3 LN:198295559    AS:hg38
@SQ     SN:chr4 LN:190214555    AS:hg38
@SQ     SN:chr5 LN:181538259    AS:hg38
@SQ     SN:chr6 LN:170805979    AS:hg38
@SQ     SN:chr7 LN:159345973    AS:hg38
@SQ     SN:chrX LN:156040895    AS:hg38
@SQ     SN:chr8 LN:145138636    AS:hg38
@SQ     SN:chr9 LN:138394717    AS:hg38
@SQ     SN:chr11        LN:135086622    AS:hg38
@SQ     SN:chr10        LN:133797422    AS:hg38
@SQ     SN:chr12        LN:133275309    AS:hg38
@SQ     SN:chr13        LN:114364328    AS:hg38
@SQ     SN:chr14        LN:107043718    AS:hg38
@SQ     SN:chr15        LN:101991189    AS:hg38
@SQ     SN:chr16        LN:90338345     AS:hg38
@SQ     SN:chr17        LN:83257441     AS:hg38
@SQ     SN:chr18        LN:80373285     AS:hg38
@SQ     SN:chr20        LN:64444167     AS:hg38
@SQ     SN:chr19        LN:58617616     AS:hg38
@SQ     SN:chrY LN:57227415     AS:hg38
@SQ     SN:chr22        LN:50818468     AS:hg38
@SQ     SN:chr21        LN:46709983     AS:hg38
@SQ     SN:chrM LN:16569        AS:hg38

The first one is my sam file and the second one gencode.v19.rRNA.interval_list

• 1 views • link
written by anjasta47 • anjasta47

As you can see there is a name mismatch for your chromosomes in those two files.

• 1 views • link
written by GenoMax GenoMax

I have no idea how I could have missed that... Thank you!

• 0 views • link
written by anjasta47 • anjasta47 
@SQ     SN:1    LN:248956422    AS:hg38
@SQ     SN:2    LN:242193529    AS:hg38
@SQ     SN:3    LN:198295559    AS:hg38
@SQ     SN:4    LN:190214555    AS:hg38
@SQ     SN:5    LN:181538259    AS:hg38
@SQ     SN:6    LN:170805979    AS:hg38
@SQ     SN:7    LN:159345973    AS:hg38
@SQ     SN:X    LN:156040895    AS:hg38
@SQ     SN:8    LN:145138636    AS:hg38
@SQ     SN:9    LN:138394717    AS:hg38
@SQ     SN:11   LN:135086622    AS:hg38
@SQ     SN:10   LN:133797422    AS:hg38
@SQ     SN:12   LN:133275309    AS:hg38
@SQ     SN:13   LN:114364328    AS:hg38
@SQ     SN:14   LN:107043718    AS:hg38
@SQ     SN:15   LN:101991189    AS:hg38
@SQ     SN:16   LN:90338345     AS:hg38
@SQ     SN:17   LN:83257441     AS:hg38
@SQ     SN:18   LN:80373285     AS:hg38
@SQ     SN:20   LN:64444167     AS:hg38
@SQ     SN:19   LN:58617616     AS:hg38
@SQ     SN:Y    LN:57227415     AS:hg38
@SQ     SN:22   LN:50818468     AS:hg38
@SQ     SN:21   LN:46709983     AS:hg38
@SQ     SN:M    LN:16569        AS:hg38

I deleted all "chr" from the gencode.v19.rRNA.interval_list file but I still get the same error. Is there something else I should get rid off or add something?

• 1 views • link
written by anjasta47 • anjasta47

M needs to become MT

• 1 views • link
written by GenoMax GenoMax

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