Question: SICER Failed to
0
gravatar for david.fear
3.7 years ago by
david.fear10
European Union
david.fear10 wrote:

I have been trying to run SICER on my ChIP-seq data set. Seequnces were aligned with bowtie2, sequencing errors and duplicates removed. The resultant .bam files were converted to .Sam and format changed to .bed. However when I try to run SICER I get an error. What am i doing wrong?

I have pasted a selection of the associated info below. Many thanks

Dave

#############################################
######           SICER v1.1            ######
#############################################
Input library directory: /tmp/tmpqLb26T
ChIP library: input_bed_file.bed
Output directory: /tmp/tmpqLb26T
Species: hg19
Threshold for redundancy allowed for reads: 1
Window size: 200 bps
Fragment size: 150 bps. The shift for reads is half of 150
Effective genome size as a fraction of the reference genome of hg19: 0.740000
Gap size: 600 bps
Evalue for identification of significant islands: 0.01
 
 
Preprocess the raw input_bed_file file to remove redundancy with threshold 1...
python /galaxy/main/deps/SICER/1.1/devteam/package_sicer_1_1/48c84c1cab2b/SICER/src/remove_redundant_reads.py -s hg19 -b /tmp/tmpqLb26T/input_bed_file.bed -t 1 -o /tmp/tmpqLb26T/input_bed_file-1-removed.bed
chr1 	Plus reads: 0 	Retained plus reads: 0 ;	Minus reads: 0 	Retained minus reads: 0
chr2 	Plus reads: 0 	Retained plus reads: 0 ;	Minus reads: 0 	Retained minus reads: 0
chr3 	Plus reads: 0 	Retained plus reads: 0 ;	Minus reads: 0 	Retained minus reads: 0
chr4 	Plus reads: 0 	Retained plus reads: 0 ;	Minus reads: 0 	Retained minus reads: 0
chr5 	Plus reads: 0 	Retained plus reads: 0 ;	Minus reads: 0 	Retained minus reads: 0
chr6 	Plus reads: 0 	Retained plus reads: 0 ;	Minus reads: 0 	Retained minus reads: 0
chr7 	Plus reads: 0 	Retained plus reads: 0 ;	Minus reads: 0 	Retained minus reads: 0
chr8 	Plus reads: 0 	Retained plus reads: 0 ;	Minus reads: 0 	Retained minus reads: 0
chr9 	Plus reads: 0 	Retained plus reads: 0 ;	Minus reads: 0 	Retained minus reads: 0
chr10 	Plus reads: 0 	Retained plus reads: 0 ;	Minus reads: 0 	Retained minus reads: 0
chr11 	Plus reads: 0 	Retained plus reads: 0 ;	Minus reads: 0 	Retained minus reads: 0
chr12 	Plus reads: 0 	Retained plus reads: 0 ;	Minus reads: 0 	Retained minus reads: 0
chr13 	Plus reads: 0 	Retained plus reads: 0 ;	Minus reads: 0 	Retained minus reads: 0

chrY 	Plus reads: 0 	Retained plus reads: 0 ;	Minus reads: 0 	Retained minus reads: 0
chrM 	Plus reads: 0 	Retained plus reads: 0 ;	Minus reads: 0 	Retained minus reads: 0
 
 
Partion the genome in windows ...
Generate summary files for input_bed_file...
python /galaxy/main/deps/SICER/1.1/devteam/package_sicer_1_1/48c84c1cab2b/SICER/src/run-make-graph-file-by-chrom.py -s hg19 -b /tmp/tmpqLb26T/input_bed_file-1-removed.bed -w 200 -i 150 -o /tmp/tmpqLb26T/input_bed_file-W200.graph
total tag count in chr1.bed is: 0.0 = 0.0+0.0
total tag count in chr2.bed is: 0.0 = 0.0+0.0
total tag count in chr3.bed is: 0.0 = 0.0+0.0
total tag count in chr4.bed is: 0.0 = 0.0+0.0
total tag count in chr5.bed is: 0.0 = 0.0+0.0
total tag count in chr6.bed is: 0.0 = 0.0+0.0

total tag count in chrX.bed is: 0.0 = 0.0+0.0
total tag count in chrY.bed is: 0.0 = 0.0+0.0
total tag count in chrM.bed is: 0.0 = 0.0+0.0


Normalize summary graph by total redundancy-removed reads per million for input_bed_file ...
python /galaxy/main/deps/SICER/1.1/devteam/package_sicer_1_1/48c84c1cab2b/SICER/src/normalize.py -i /tmp/tmpqLb26T/input_bed_file-W200.graph -a 3 -t 1000000 -o /tmp/tmpqLb26T/input_bed_file-W200-normalized.graph


Convert the normalized summary graph into wig vstep format...
sh /galaxy/main/deps/SICER/1.1/devteam/package_sicer_1_1/48c84c1cab2b/SICER/src/variableStep.sh /tmp/tmpqLb26T/input_bed_file-W200-normalized.graph /tmp/tmpqLb26T/input_bed_file-W200-normalized.wig input_bed_file 200
 
 
Find significant islands with E-value 0.01 for input_bed_file...
python /galaxy/main/deps/SICER/1.1/devteam/package_sicer_1_1/48c84c1cab2b/SICER/src/find_islands_in_pr.py -s hg19 -b /tmp/tmpqLb26T/input_bed_file-W200.graph -w 200 -g 600 -t 0.740000 -e 0.01 -f /tmp/tmpqLb26T/input_bed_file-W200-G600-E0.01.scoreisland
Species:  hg19
Window_size:  200
Gap size:  600
E value is: 0.01
Total read count: 0.0
Genome Length:  3095693983
Effective genome Length:  2290813547
Window average: 0.0
Window pvalue: 0.2


Filter reads with identified significant islands for input_bed_file...
python /galaxy/main/deps/SICER/1.1/devteam/package_sicer_1_1/48c84c1cab2b/SICER/utility/filter_raw_tags_by_islands.py -s hg19 -a /tmp/tmpqLb26T/input_bed_file-1-removed.bed -i 150 -b /tmp/tmpqLb26T/input_bed_file-W200-G600-E0.01.scoreisland  -o  /tmp/tmpqLb26T/input_bed_file-W200-G600-E0.01-islandfiltered.bed


Make summary graph with filtered reads for input_bed_file...
python /galaxy/main/deps/SICER/1.1/devteam/package_sicer_1_1/48c84c1cab2b/SICER/src/run-make-graph-file-by-chrom.py -s hg19 -b /tmp/tmpqLb26T/input_bed_file-W200-G600-E0.01-islandfiltered.bed -w 200 -i 150 -o /tmp/tmpqLb26T/input_bed_file-W200-G600-E0.01-islandfiltered.graph
total tag count in chr1.bed is: 0.0 = 0.0+0.0
total tag count in chr2.bed is: 0.0 = 0.0+0.0
total tag count in chr3.bed is: 0.0 = 0.0+0.0
total tag count in chr4.bed is: 0.0 = 0.0+0.0

total tag count in chrX.bed is: 0.0 = 0.0+0.0
total tag count in chrY.bed is: 0.0 = 0.0+0.0
total tag count in chrM.bed is: 0.0 = 0.0+0.0


Normalize summary graph with filtered reads for input_bed_file by total island filtered reads per million...
python /galaxy/main/deps/SICER/1.1/devteam/package_sicer_1_1/48c84c1cab2b/SICER/src/normalize.py -i /tmp/tmpqLb26T/input_bed_file-W200-G600-E0.01-islandfiltered.graph -a 3 -t 1000000 -o /tmp/tmpqLb26T/input_bed_file-W200-G600-E0.01-islandfiltered-normalized.graph


Convert the summary graph made with the filtered reads into wig vstep format and normalize by total island-filtered read count per million...
sh /galaxy/main/deps/SICER/1.1/devteam/package_sicer_1_1/48c84c1cab2b/SICER/src/variableStep.sh /tmp/tmpqLb26T/input_bed_file-W200-G600-E0.01-islandfiltered-normalized.graph /tmp/tmpqLb26T/input_bed_file-W200-G600-E0.01-islandfiltered-normalized.wig input_bed_file 200


Done!

Additionally, these warnings were reported:
Warning: chr1 reads do not exist in /tmp/tmpqLb26T/input_bed_file.bed
Warning: chr2 reads do not exist in /tmp/tmpqLb26T/input_bed_file.bed
Warning: chr3 reads do not exist in /tmp/tmpqLb26T/input_bed_file.bed
Warning: chr4 reads do not exist in /tmp/tmpqLb26T/input_bed_file.bed

Warning: chr21 reads do not exist in /tmp/tmpqLb26T/input_bed_file.bed
Warning: chr22 reads do not exist in /tmp/tmpqLb26T/input_bed_file.bed
Warning: chrX reads do not exist in /tmp/tmpqLb26T/input_bed_file.bed
Warning: chrY reads do not exist in /tmp/tmpqLb26T/input_bed_file.bed
Warning: chrM reads do not exist in /tmp/tmpqLb26T/input_bed_file.bed
Warning: chr1 reads do not exist in /tmp/tmpqLb26T/input_bed_file-1-removed.bed
Warning: chr2 reads do not exist in /tmp/tmpqLb26T/input_bed_file-1-removed.bed
Warning: chr3 reads do not exist in /tmp/tmpqLb26T/input_bed_file-1-removed.bed
Warning: chr4 reads do not exist in /tmp/tmpqLb26T/input_bed_file-1-removed.bed
Warning: chr5 reads do not exist in /tmp/tmpqLb26T/input_bed_file-1-removed.bed

Warning: chrX reads do not exist in /tmp/tmpqLb26T/input_bed_file-1-removed.bed
Warning: chrY reads do not exist in /tmp/tmpqLb26T/input_bed_file-1-removed.bed
Warning: chrM reads do not exist in /tmp/tmpqLb26T/input_bed_file-1-removed.bed
Traceback (most recent call last):
  File "/galaxy/main/deps/SICER/1.1/devteam/package_sicer_1_1/48c84c1cab2b/SICER/src/find_islands_in_pr.py", line 270, in <module>
    main(sys.argv)
  File "/galaxy/main/deps/SICER/1.1/devteam/package_sicer_1_1/48c84c1cab2b/SICER/src/find_islands_in_pr.py", line 204, in main
    background = Background_island_probscore_statistics.Background_island_probscore_statistics(total_read_count, opt.window_size, opt.gap, window_pvalue, genome_length, bin_size);
  File "/galaxy/main/deps/SICER/1.1/devteam/package_sicer_1_1/48c84c1cab2b/SICER/lib/Background_island_probscore_statistics.py", line 45, in __init__
    prob = self.poisson(index, self.average);
  File "/galaxy/main/deps/SICER/1.1/devteam/package_sicer_1_1/48c84c1cab2b/SICER/lib/Background_island_probscore_statistics.py", line 126, in poisson
    exponent = -average + i*log(average) - self.factln(i);
ValueError: math domain error
Traceback (most recent call last):
  File "/galaxy/main/deps/SICER/1.1/devteam/package_sicer_1_1/48c84c1cab2b/SICER/utility/filter_raw_tags_by_islands.py", line 111, in <module>
    main(sys.argv)
  File "/galaxy/main/deps/SICER/1.1/devteam/package_sicer_1_1/48c84c1cab2b/SICER/utility/filter_raw_tags_by_islands.py", line 99, in main
    islands = BED.BED(opt.species, opt.islandbedfile, "BED3", 0);
  File "/galaxy/main/deps/SICER/1.1/devteam/package_sicer_1_1/48c84c1cab2b/SICER/lib/BED.py", line 202, in __init__
    infile = open(file);
IOError: [Errno 2] No such file or directory: '/tmp/tmpqLb26T/input_bed_file-W200-G600-E0.01.scoreisland'
grep: /tmp/tmpqLb26T/input_bed_file-W200-G600-E0.01-islandfiltered.bed: No such file or directory
Warning: chr1 reads do not exist in /tmp/tmpqLb26T/input_bed_file-W200-G600-E0.01-islandfiltered.bed
grep: /tmp/tmpqLb26T/input_bed_file-W200-G600-E0.01-islandfiltered.bed: No such file or directory
Warning: chr2 reads do not exist in /tmp/tmpqLb26T/input_bed_file-W200-G600-E0.01-islandfiltered.bed
grep: /tmp/tmpqLb26T/input_bed_file-W200-G600-E0.01-islandfiltered.bed: No such file or directory
Warning: chr3 reads do not exist in /tmp/tmpqLb26T/input_bed_file-W200-G600-E0.01-islandfiltered.bed
grep: /tmp/tmpqLb26T/input_bed_file-W200-G600-E0.01-islandfiltered.bed: No such file or directory
Warning: chr4 reads do not exist in /tmp/tmpqLb26T/input_bed_file-W200-G600-E0.01-islandfiltered.bed
grep: /tmp/tmpqLb26T/input_bed_file-W200-G600-E0.01-islandfiltered.bed: No such file or directory
Warning: chr5 reads do not exist in /tmp/tmpqLb26T/input_bed_file-W200-G600-E0.01-islandfiltered.bed
grep: /tmp/tmpqLb26T/input_bed_file-W200-G600-E0.01-islandfiltered.bed: No such file or directory
Warning: chr6 reads do not exist in /tmp/tmpqLb26T/input_bed_file-W200-G600-E0.01-islandfiltered.bed
grep: /tmp/tmpqLb26T/input_bed_file-W200-G600-E0.01-islandfiltered.bed: No such file or directory
Warning: chr7 reads do not exist in /tmp/tmpqLb26T/input_bed_file-W200-G600-E0.01-islandfiltered.bed
grep: /tmp/tmpqLb26T/input_bed_file-W200-G600-E0.01-islandfiltered.bed: No such file or directory
Warning: chr8 reads do not exist in /tmp/tmpqLb26T/input_bed_file-W200-G600-E0.01-islandfiltered.bed
grep: /tmp/tmpqLb26T/input_bed_file-W200-G600-E0.01-islandfiltered.bed: No such file or directory
Warning: chr9 reads do not exist in /tmp/tmpqLb26T/input_bed_file-W200-G600-E0.01-islandfiltered.bed
grep: /tmp/tmpqLb26T/input_bed_file-W200-G600-E0.01-islandfiltered.bed: No such file or directory
Warning: chr10 reads do not exist in /tmp/tmpqLb26T/input_bed_file-W200-G600-E0.01-islandfiltered.bed
grep: /tmp/tmpqLb26T/input_bed_file-W200-G600-E0.01-islandfiltered.bed: No such file or directory
Warning: chr11 reads do not exist in /tmp/tmpqLb26T/input_bed_file-W200-G600-E0.01-islandfiltered.bed
grep: /tmp/tmpqLb26T/input_bed_file-W200-G600-E0.01-islandfiltered.bed: No such file or directory
Warning: chr12 reads do not exist in /tmp/tmpqLb26T/input_bed_file-W200-G600-E0.01-islandfiltered.bed
grep: /tmp/tmpqLb26T/input_bed_file-W200-G600-E0.01-islandfiltered.bed: No such file or directory
Warning: chr13 reads do not exist in /tmp/tmpqLb26T/input_bed_file-W200-G600-E0.01-islandfiltered.bed
grep: /tmp/tmpqLb26T/input_bed_file-W200-G600-E0.01-islandfiltered.bed: No such file or directory
Warning: chr14 reads do not exist in /tmp/tmpqLb26T/input_bed_file-W200-G600-E0.01-islandfiltered.bed
grep: /tmp/tmpqLb26T/input_bed_file-W200-G600-E0.01-islandfiltered.bed: No such file or directory
Warning: chr15 reads do not exist in /tmp/tmpqLb26T/input_bed_file-W200-G600-E0.01-islandfiltered.bed
grep: /tmp/tmpqLb26T/input_bed_file-W200-G600-E0.01-islandfiltered.bed: No such file or directory
Warning: chr16 reads do not exist in /tmp/tmpqLb26T/input_bed_file-W200-G600-E0.01-islandfiltered.bed
grep: /tmp/tmpqLb26T/input_bed_file-W200-G600-E0.01-islandfiltered.bed: No such file or directory
Warning: chr17 reads do not exist in /tmp/tmpqLb26T/input_bed_file-W200-G600-E0.01-islandfiltered.bed
grep: /tmp/tmpqLb26T/input_bed_file-W200-G600-E0.01-islandfiltered.bed: No such file or directory
Warning: chr18 reads do not exist in /tmp/tmpqLb26T/input_bed_file-W200-G600-E0.01-islandfiltered.bed
grep: /tmp/tmpqLb26T/input_bed_file-W200-G600-E0.01-islandfiltered.bed: No such file or directory
Warning: chr19 reads do not exist in /tmp/tmpqLb26T/input_bed_file-W200-G600-E0.01-islandfiltered.bed
grep: /tmp/tmpqLb26T/input_bed_file-W200-G600-E0.01-islandfiltered.bed: No such file or directory
Warning: chr20 reads do not exist in /tmp/tmpqLb26T/input_bed_file-W200-G600-E0.01-islandfiltered.bed
grep: /tmp/tmpqLb26T/input_bed_file-W200-G600-E0.01-islandfiltered.bed: No such file or directory
Warning: chr21 reads do not exist in /tmp/tmpqLb26T/input_bed_file-W200-G600-E0.01-islandfiltered.bed
grep: /tmp/tmpqLb26T/input_bed_file-W200-G600-E0.01-islandfiltered.bed: No such file or directory
Warning: chr22 reads do not exist in /tmp/tmpqLb26T/input_bed_file-W200-G600-E0.01-islandfiltered.bed
grep: /tmp/tmpqLb26T/input_bed_file-W200-G600-E0.01-islandfiltered.bed: No such file or directory
Warning: chrX reads do not exist in /tmp/tmpqLb26T/input_bed_file-W200-G600-E0.01-islandfiltered.bed
grep: /tmp/tmpqLb26T/input_bed_file-W200-G600-E0.01-islandfiltered.bed: No such file or directory
Warning: chrY reads do not exist in /tmp/tmpqLb26T/input_bed_file-W200-G600-E0.01-islandfiltered.bed
grep: /tmp/tmpqLb26T/input_bed_file-W200-G600-E0.01-islandfiltered.bed: No such file or directory
Warning: chrM reads do not exist in /tmp/tmpqLb26T/input_bed_file-W200-G600-E0.01-islandfiltered.bed
chip-seq • 1.6k views
ADD COMMENTlink modified 3.7 years ago by Jennifer Hillman Jackson23k • written 3.7 years ago by david.fear10
0
gravatar for Jennifer Hillman Jackson
3.7 years ago by
United States
Jennifer Hillman Jackson23k wrote:

Hello,

Have you checked the input file? Does it have hits in it?

If it does, then check the chromosome identifiers for a mismatch. If you didn't run Bowtie2 in Galaxy against hg19, then you used UCSC's version of hg19 externally? Sourced from Galaxy's rsync or UCSC downloads?
https://wiki.galaxyproject.org/Support#Reference_genomes

You will probably want to adjust other default options like fragment size to match your data (as needed) for the best results, but that is secondary parameter tuning and shouldn't cause this type of error that I am aware of.

Thanks, Jen, Galaxy team

ADD COMMENTlink written 3.7 years ago by Jennifer Hillman Jackson23k
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