ERROR: no associated .bai file found with bam.

[Former user]

Hi, I am trying to run ROSE on some H3K27Ac peak sets but am getting an error stating there is no bam index (when there is).

ls *bam*

E3381K27WT-H3K27Ac.dedupe.bam      E3387K27WT-H3K27Ac.dedupe.bam      ROSE_bamToGFF.py
E3381K27WT-H3K27Ac.dedupe.bam.bai  E3387K27WT-H3K27Ac.dedupe.bam.bai  ROSE_bamToGFF_turbo.py
E3381K27WT-Input.dedupe.bam        E3387K27WT-Input.dedupe.bam
E3381K27WT-Input.dedupe.bam.bai    E3387K27WT-Input.dedupe.bam.bai

The script seems to be continuing but ROSE_bamToGFF is throwing these errors.

python ROSE_main.py -i Wt_H3K27Ac_peaks_filtered_intersected.gff -r E3381K27WT-H3K27Ac.dedupe.bam -o WT_AC_ROSE -g HG38 -b E3387K27WT-H3K27Ac.dedupe.bam -c E3387K27WT_Input.dedupe.bam -t 2500
USING Wt_H3K27Ac_peaks_filtered_intersected.gff AS THE INPUT GFF
USING HG38 AS THE GENOME
MAKING START DICT
LOADING IN GFF REGIONS
CHECKING INPUT TO MAKE SURE EACH REGION HAS A UNIQUE IDENTIFIER
REFERENCE COLLECTION PASSES QC
STITCHING REGIONS TOGETHER
PERFORMING REGION STITCHING
REMOVED 8321 LOCI BECAUSE THEY WERE CONTAINED BY A TSS
REMOVED 92 STITCHED LOCI BECAUSE THEY OVERLAPPED MULTIPLE TSSs
ADDED BACK 242 ORIGINAL LOCI
MAKING GFF FROM STITCHED COLLECTION
WRITING STITCHED GFF TO DISK AS WT_AC_ROSE/gff/Wt_H3K27Ac_peaks_filtered_intersected_12KB_STITCHED_TSS_DISTAL.gff
OUTPUT WILL BE WRITTEN TO  WT_AC_ROSE/Wt_H3K27Ac_peaks_filtered_intersected_12KB_STITCHED_TSS_DISTAL_ENHANCER_REGION_MAP.txt
python ROSE_bamToGFF.py -f 1 -e 200 -r -m 1 -b E3381K27WT-H3K27Ac.dedupe.bam -i WT_AC_ROSE/gff/Wt_H3K27Ac_peaks_filtered_intersected_12KB_STITCHED_TSS_DISTAL.gff -o WT_AC_ROSE/mappedGFF/Wt_H3K27Ac_peaks_filtered_intersected_12KB_STITCHED_TSS_DISTAL_E3381K27WT-H3K27Ac.dedupe.bam_MAPPED.gff &
python ROSE_bamToGFF.py -f 1 -e 200 -r -m 1 -b E3381K27WT-H3K27Ac.dedupe.bam -i Wt_H3K27Ac_peaks_filtered_intersected.gff -o WT_AC_ROSE/mappedGFF/Wt_H3K27Ac_peaks_filtered_intersected_E3381K27WT-H3K27Ac.dedupe.bam_MAPPED.gff &
python ROSE_bamToGFF.py -f 1 -e 200 -r -m 1 -b E3387K27WT_Input.dedupe.bam -i WT_AC_ROSE/gff/Wt_H3K27Ac_peaks_filtered_intersected_12KB_STITCHED_TSS_DISTAL.gff -o WT_AC_ROSE/mappedGFF/Wt_H3K27Ac_peaks_filtered_intersected_12KB_STITCHED_TSS_DISTAL_E3387K27WT_Input.dedupe.bam_MAPPED.gff &
{'matrix': '1', 'extension': '200', 'floor': '1', 'sense': 'both', 'output': 'WT_AC_ROSE/mappedGFF/Wt_H3K27Ac_peaks_filtered_intersected_12KB_STITCHED_TSS_DISTAL_E3381K27WT-H3K27Ac.dedupe.bam_MAPPED.gff', 'bam': 'E3381K27WT-H3K27Ac.dedupe.bam', 'rpm': True, 'input': 'WT_AC_ROSE/gff/Wt_H3K27Ac_peaks_filtered_intersected_12KB_STITCHED_TSS_DISTAL.gff'}
[]
mapping to GFF and making a matrix with fixed bin number
python ROSE_bamToGFF.py -f 1 -e 200 -r -m 1 -b E3387K27WT_Input.dedupe.bam -i Wt_H3K27Ac_peaks_filtered_intersected.gff -o WT_AC_ROSE/mappedGFF/Wt_H3K27Ac_peaks_filtered_intersected_E3387K27WT_Input.dedupe.bam_MAPPED.gff &
{'matrix': '1', 'extension': '200', 'floor': '1', 'sense': 'both', 'output': 'WT_AC_ROSE/mappedGFF/Wt_H3K27Ac_peaks_filtered_intersected_E3381K27WT-H3K27Ac.dedupe.bam_MAPPED.gff', 'bam': 'E3381K27WT-H3K27Ac.dedupe.bam', 'rpm': True, 'input': 'Wt_H3K27Ac_peaks_filtered_intersected.gff'}
[]
mapping to GFF and making a matrix with fixed bin number
python ROSE_bamToGFF.py -f 1 -e 200 -r -m 1 -b E3387K27WT-H3K27Ac.dedupe.bam -i WT_AC_ROSE/gff/Wt_H3K27Ac_peaks_filtered_intersected_12KB_STITCHED_TSS_DISTAL.gff -o WT_AC_ROSE/mappedGFF/Wt_H3K27Ac_peaks_filtered_intersected_12KB_STITCHED_TSS_DISTAL_E3387K27WT-H3K27Ac.dedupe.bam_MAPPED.gff &
{'matrix': '1', 'extension': '200', 'floor': '1', 'sense': 'both', 'output': 'WT_AC_ROSE/mappedGFF/Wt_H3K27Ac_peaks_filtered_intersected_12KB_STITCHED_TSS_DISTAL_E3387K27WT_Input.dedupe.bam_MAPPED.gff', 'bam': 'E3387K27WT_Input.dedupe.bam', 'rpm': True, 'input': 'WT_AC_ROSE/gff/Wt_H3K27Ac_peaks_filtered_intersected_12KB_STITCHED_TSS_DISTAL.gff'}
[]
ERROR: no associated .bai file found with bam. Must use a sorted bam with accompanying index file
Usage: ROSE_bamToGFF.py [options] -b [SORTED BAMFILE] -i [INPUTFILE] -o [OUTPUTFILE]

Options:
  -h, --help            show this help message and exit
  -b BAM, --bam=BAM     Enter .bam file to be processed.
  -i INPUT, --input=INPUT
                        Enter .gff or ENRICHED REGION file to be processed.
  -o OUTPUT, --output=OUTPUT
                        Enter the output filename.
  -s SENSE, --sense=SENSE
                        Map to '+','-' or 'both' strands. Default maps to
                        both.
  -f FLOOR, --floor=FLOOR
                        Sets a read floor threshold necessary to count towards
                        density
  -e EXTENSION, --extension=EXTENSION
                        Extends reads by n bp. Default value is 200bp
  -r, --rpm             Normalizes density to reads per million (rpm)
  -m MATRIX, --matrix=MATRIX
                        Outputs a variable bin sized matrix. User must specify
                        number of bins.
python ROSE_bamToGFF.py -f 1 -e 200 -r -m 1 -b E3387K27WT-H3K27Ac.dedupe.bam -i Wt_H3K27Ac_peaks_filtered_intersected.gff -o WT_AC_ROSE/mappedGFF/Wt_H3K27Ac_peaks_filtered_intersected_E3387K27WT-H3K27Ac.dedupe.bam_MAPPED.gff &
{'matrix': '1', 'extension': '200', 'floor': '1', 'sense': 'both', 'output': 'WT_AC_ROSE/mappedGFF/Wt_H3K27Ac_peaks_filtered_intersected_E3387K27WT_Input.dedupe.bam_MAPPED.gff', 'bam': 'E3387K27WT_Input.dedupe.bam', 'rpm': True, 'input': 'Wt_H3K27Ac_peaks_filtered_intersected.gff'}
[]
ERROR: no associated .bai file found with bam. Must use a sorted bam with accompanying index file
Usage: ROSE_bamToGFF.py [options] -b [SORTED BAMFILE] -i [INPUTFILE] -o [OUTPUTFILE]

Options:
  -h, --help            show this help message and exit
  -b BAM, --bam=BAM     Enter .bam file to be processed.
  -i INPUT, --input=INPUT
                        Enter .gff or ENRICHED REGION file to be processed.
  -o OUTPUT, --output=OUTPUT
                        Enter the output filename.
  -s SENSE, --sense=SENSE
                        Map to '+','-' or 'both' strands. Default maps to
                        both.
  -f FLOOR, --floor=FLOOR
                        Sets a read floor threshold necessary to count towards
                        density
  -e EXTENSION, --extension=EXTENSION
                        Extends reads by n bp. Default value is 200bp
  -r, --rpm             Normalizes density to reads per million (rpm)
  -m MATRIX, --matrix=MATRIX
                        Outputs a variable bin sized matrix. User must specify
                        number of bins.
{'matrix': '1', 'extension': '200', 'floor': '1', 'sense': 'both', 'output': 'WT_AC_ROSE/mappedGFF/Wt_H3K27Ac_peaks_filtered_intersected_12KB_STITCHED_TSS_DISTAL_E3387K27WT-H3K27Ac.dedupe.bam_MAPPED.gff', 'bam': 'E3387K27WT-H3K27Ac.dedupe.bam', 'rpm': True, 'input': 'WT_AC_ROSE/gff/Wt_H3K27Ac_peaks_filtered_intersected_12KB_STITCHED_TSS_DISTAL.gff'}
[]
mapping to GFF and making a matrix with fixed bin number
PAUSING TO MAP
{'matrix': '1', 'extension': '200', 'floor': '1', 'sense': 'both', 'output': 'WT_AC_ROSE/mappedGFF/Wt_H3K27Ac_peaks_filtered_intersected_E3387K27WT-H3K27Ac.dedupe.bam_MAPPED.gff', 'bam': 'E3387K27WT-H3K27Ac.dedupe.bam', 'rpm': True, 'input': 'Wt_H3K27Ac_peaks_filtered_intersected.gff'}
[]
mapping to GFF and making a matrix with fixed bin number
using a MMR value of 10.1709
has chr
Number lines processed
0
using a MMR value of 10.1709
has chr
Number lines processed
0
using a MMR value of 11.1455
has chr
Number lines processed
0
using a MMR value of 11.1455
has chr
Number lines processed
0
WAITING FOR MAPPING TO COMPLETE. ELAPSED TIME (MIN):

[cmonger]

Solved this issue by putting the files in the same directory structure as in the example data and calling the bam file path with ./

E.G:

python ROSE_main.py -g HG38 -i ./data/Wt_H3K27Ac_peaks_filtered_intersected.gff -r ./data/E3381K27WT-H3K27Ac.dedupe.sorted.bam -c ./data/E3381K27WT-Input.dedupe.sorted.bam -o WT_AC_ROSE/ -s 12500 -t 2500

This program is very finicky with how it is called….

I think a small but hugely beneficial improvement to the program would be to put in a check at the start of ROSE_main.py to check that all input files exist and are findable, and to kill the script before executing the subscripts if not.