[miso-users] Error: tagBam call failed when running pe_utils.py --compute-insert-len

[Gu Mi]

Dear All:

I am using MISO to test for differential exon usage between a control and a
treatment group. I got an error *when computing the insert length
distribution using pe_utils.py --compute-insert-len*. I list the steps I
used below:

1. sort the BAM file from TopHat (by coordinate):
samtools sort control.bam control_sorted

2. index the BAM file:
samtools index control_sorted.bam control_sorted.bai

3. run pe_utils.py:
python pe_utils.py --compute-insert-len controlam
/directories/exons/Homo_sapiens.GRCh37.65.min_1000.const_exons.gff
--output-dir /directories/insert-dist/

After the command above, I got the* error message*:

Preparing to call bedtools 'tagBam'
tagBam -i control.bam -files
/directories/exons/Homo_sapiens.GRCh37.65.min_1000.const_exons.gff -labels
gff -intervals -f 1 | samtools view - -h | egrep '^@|:gff:' | samtools view
- -Shb -o
/directories/insert-dist/bam2gff_Homo_sapiens.GRCh37.65.min_1000.const_exons.gff/control.bam
[samopen] SAM header is present: 25 sequences.
[sam_read1] reference 'ID:TopHat
 CL:/informatics/tools/Linux-AS5/bin/tophat -o Lane3 -g 1 --coverage-search
--microexon -r 100 --phred64-quals --library-type fr-unstranded -p 4 -G
gene_models/Homo_sapiens.GRCh37.72_norm.gtf
--transcriptome-index=gene_models/transcripts
/directories/Genomes/NCBI_Jul-09-2012/Human/bowtie/human_ref_genome
Lane3_1.fq.gz Lane3_2.fq.gz     VN:1.4.1
' is recognized as '*'.
[main_samview] truncated file.
Traceback (most recent call last):
  File "/pe_utils.py", line 520, in <module>
    main()
  File "pe_utils.py", line 517, in main
    sd_max=sd_max)
  File "pe_utils.py", line 271, in compute_insert_len
    output_dir)
  File "exon_utils.py", line 185, in map_bam2gff
    raise Exception, "Error: tagBam call failed."
Exception: Error: tagBam call failed.

I used *Homo_sapiens.GRCh37.72_norm.gtf* from Ensembl as the annotation
file when preparing my data, but downloaded

   - Human genome (hg19) alternative events
v2.0<http://genes.mit.edu/burgelab/miso/annotations/ver2/miso_annotations_hg19_v2.zip>

from the MISO website and unzipped. I saw it is based
on Homo_sapiens.GRCh37.65. Is this the version problem? If so, could anyone
provide the latest GFF3 file for use? Thank you for your suggestions!

Best,
Gu
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