Good number of reads but very few peaks
Abhishek Singh
samtools view -F 0x904 -c p300.sort.bam
16991725
However, when I call peaks using macs2
macs2 callpeak -t p300.sort.bam -c input.bam -n p300.NOF --gsize hs
INFO @ Fri, 03 Nov 2017 14:35:00:
# Command line: callpeak -t p300.sort.bam -c input.bam -n p300.NOF --gsize hs
# ARGUMENTS LIST:
# name = p300.NOF
# format = AUTO
# ChIP-seq file = ['p300.sort.bam']
# control file = ['input.bam']
# effective genome size = 2.70e+09
# band width = 300
# model fold = [5, 50]
# qvalue cutoff = 5.00e-02
# Larger dataset will be scaled towards smaller dataset.
# Range for calculating regional lambda is: 1000 bps and 10000 bps
# Broad region calling is off
# Paired-End mode is off
INFO @ Fri, 03 Nov 2017 14:35:00: #1 read tag files...
INFO @ Fri, 03 Nov 2017 14:35:00: #1 read treatment tags...
INFO @ Fri, 03 Nov 2017 14:35:00: Detected format is: BAM
INFO @ Fri, 03 Nov 2017 14:35:00: * Input file is gzipped.
INFO @ Fri, 03 Nov 2017 14:35:03: 1000000
INFO @ Fri, 03 Nov 2017 14:35:06: 2000000
INFO @ Fri, 03 Nov 2017 14:35:08: 3000000
INFO @ Fri, 03 Nov 2017 14:35:11: 4000000
INFO @ Fri, 03 Nov 2017 14:35:14: 5000000
INFO @ Fri, 03 Nov 2017 14:35:16: 6000000
INFO @ Fri, 03 Nov 2017 14:35:19: 7000000
INFO @ Fri, 03 Nov 2017 14:35:22: 8000000
INFO @ Fri, 03 Nov 2017 14:35:24: 9000000
INFO @ Fri, 03 Nov 2017 14:35:27: 10000000
INFO @ Fri, 03 Nov 2017 14:35:30: 11000000
INFO @ Fri, 03 Nov 2017 14:35:32: 12000000
INFO @ Fri, 03 Nov 2017 14:35:35: 13000000
INFO @ Fri, 03 Nov 2017 14:35:38: 14000000
INFO @ Fri, 03 Nov 2017 14:35:40: 15000000
INFO @ Fri, 03 Nov 2017 14:35:43: 16000000
INFO @ Fri, 03 Nov 2017 14:35:46: #1.2 read input tags...
INFO @ Fri, 03 Nov 2017 14:35:46: Detected format is: BAM
INFO @ Fri, 03 Nov 2017 14:35:46: * Input file is gzipped.
INFO @ Fri, 03 Nov 2017 14:35:48: 1000000
INFO @ Fri, 03 Nov 2017 14:35:50: 2000000
INFO @ Fri, 03 Nov 2017 14:35:52: 3000000
INFO @ Fri, 03 Nov 2017 14:35:54: 4000000
INFO @ Fri, 03 Nov 2017 14:35:57: 5000000
INFO @ Fri, 03 Nov 2017 14:35:59: 6000000
INFO @ Fri, 03 Nov 2017 14:36:01: 7000000
INFO @ Fri, 03 Nov 2017 14:36:03: 8000000
INFO @ Fri, 03 Nov 2017 14:36:06: 9000000
INFO @ Fri, 03 Nov 2017 14:36:08: 10000000
INFO @ Fri, 03 Nov 2017 14:36:10: 11000000
INFO @ Fri, 03 Nov 2017 14:36:12: 12000000
INFO @ Fri, 03 Nov 2017 14:36:14: 13000000
INFO @ Fri, 03 Nov 2017 14:36:17: 14000000
INFO @ Fri, 03 Nov 2017 14:36:19: 15000000
INFO @ Fri, 03 Nov 2017 14:36:21: 16000000
INFO @ Fri, 03 Nov 2017 14:36:23: 17000000
INFO @ Fri, 03 Nov 2017 14:36:25: 18000000
INFO @ Fri, 03 Nov 2017 14:36:27: 19000000
INFO @ Fri, 03 Nov 2017 14:36:30: 20000000
INFO @ Fri, 03 Nov 2017 14:36:32: 21000000
INFO @ Fri, 03 Nov 2017 14:36:34: 22000000
INFO @ Fri, 03 Nov 2017 14:36:36: 23000000
INFO @ Fri, 03 Nov 2017 14:36:39: 24000000
INFO @ Fri, 03 Nov 2017 14:36:41: 25000000
INFO @ Fri, 03 Nov 2017 14:36:43: 26000000
INFO @ Fri, 03 Nov 2017 14:36:45: 27000000
INFO @ Fri, 03 Nov 2017 14:36:47: 28000000
INFO @ Fri, 03 Nov 2017 14:36:48: 29000000
INFO @ Fri, 03 Nov 2017 14:36:50: 30000000
INFO @ Fri, 03 Nov 2017 14:36:51: 31000000
INFO @ Fri, 03 Nov 2017 14:36:52: 32000000
INFO @ Fri, 03 Nov 2017 14:36:53: 33000000
INFO @ Fri, 03 Nov 2017 14:36:55: 34000000
INFO @ Fri, 03 Nov 2017 14:36:56: 35000000
INFO @ Fri, 03 Nov 2017 14:36:57: #1 tag size is determined as 65 bps
INFO @ Fri, 03 Nov 2017 14:36:57: #1 tag size = 65
INFO @ Fri, 03 Nov 2017 14:36:57: #1 total tags in treatment: 16991725
INFO @ Fri, 03 Nov 2017 14:36:57: #1 user defined the maximum tags...
INFO @ Fri, 03 Nov 2017 14:36:57: #1 filter out redundant tags at the same location and the same strand by allowing at most 1 tag(s)
INFO @ Fri, 03 Nov 2017 14:36:57: #1 tags after filtering in treatment: 16693135
INFO @ Fri, 03 Nov 2017 14:36:57: #1 Redundant rate of treatment: 0.02
INFO @ Fri, 03 Nov 2017 14:36:57: #1 total tags in control: 27749309
INFO @ Fri, 03 Nov 2017 14:36:57: #1 user defined the maximum tags...
INFO @ Fri, 03 Nov 2017 14:36:57: #1 filter out redundant tags at the same location and the same strand by allowing at most 1 tag(s)
INFO @ Fri, 03 Nov 2017 14:36:58: #1 tags after filtering in control: 26326831
INFO @ Fri, 03 Nov 2017 14:36:58: #1 Redundant rate of control: 0.05
INFO @ Fri, 03 Nov 2017 14:36:58: #1 finished!
INFO @ Fri, 03 Nov 2017 14:36:58: #2 Build Peak Model...
INFO @ Fri, 03 Nov 2017 14:36:58: #2 looking for paired plus/minus strand peaks...
INFO @ Fri, 03 Nov 2017 14:36:59: #2 number of paired peaks: 1218
INFO @ Fri, 03 Nov 2017 14:36:59: start model_add_line...
INFO @ Fri, 03 Nov 2017 14:36:59: start X-correlation...
INFO @ Fri, 03 Nov 2017 14:36:59: end of X-cor
INFO @ Fri, 03 Nov 2017 14:36:59: #2 finished!
INFO @ Fri, 03 Nov 2017 14:36:59: #2 predicted fragment length is 62 bps
INFO @ Fri, 03 Nov 2017 14:36:59: #2 alternative fragment length(s) may be 62,163,194,283,441,510 bps
INFO @ Fri, 03 Nov 2017 14:36:59: #2.2 Generate R script for model : p300.NOF_model.r
WARNING @ Fri, 03 Nov 2017 14:36:59: #2 Since the d (62) calculated from paired-peaks are smaller than 2*tag length, it may be influenced by unknown sequencing problem!
WARNING @ Fri, 03 Nov 2017 14:36:59: #2 You may need to consider one of the other alternative d(s): 62,163,194,283,441,510
WARNING @ Fri, 03 Nov 2017 14:36:59: #2 You can restart the process with --nomodel --extsize XXX with your choice or an arbitrary number. Nontheless, MACS will continute computing.
INFO @ Fri, 03 Nov 2017 14:36:59: #3 Call peaks...
INFO @ Fri, 03 Nov 2017 14:36:59: #3 Pre-compute pvalue-qvalue table...
INFO @ Fri, 03 Nov 2017 14:38:13: #3 Call peaks for each chromosome...
INFO @ Fri, 03 Nov 2017 14:38:52: #4 Write output xls file... p300.NOF_peaks.xls
INFO @ Fri, 03 Nov 2017 14:38:52: #4 Write peak in narrowPeak format file... p300.NOF_peaks.narrowPeak
INFO @ Fri, 03 Nov 2017 14:38:52: #4 Write summits bed file... p300.NOF_summits.bed
INFO @ Fri, 03 Nov 2017 14:38:52: Done!
I did try putting up different --extsize with option --nomodel but it didn't work.
Can anyone suggest the reason behind this strange behaviour. I have never seen such a thing.
Thank you
Ian Donaldson
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John Urban
Abhishek Singh
Are you expecting point source peaks? or broad enrichment zones?
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Moshe Olshansky
We also have an old P300 ChIP-Seq (mouse) for which we could not do anything. And yes, very low redundancy rate in the treatment sample may suggest that your reads are spread all over the genome. If I remember correctly we had a similar problem.