Error in smooth.spline

[evaham1]

Hello,

Thanks for the great tool, I have run into the following error which means Fithichip is not able to compute the statistical significance of input interactions. I am running Fithichip through Singularity and have provided HiCPro valid pairs as my input.

Any advice would be really appreciated!

Here is the output of my run:

/flask/scratch/briscoej/hamrude/atac_neural_plate_border/NF-hichip/run_fithichip/data/configfile_NF_r1

 ================ Verifying input configuration parameters =================
***** Specified output directory : /flask/scratch/briscoej/hamrude/atac_neural_plate_border/output/NF-hichip/NF_FitHiChiP_r1
====>>> User did not provide any file in the COOL= option or the HIC= option or the Bed= option of the configuration file
==>>> So, the input interaction file is from the HiC-pro (valid pairs format)
==>>> But the matrix and bin interval files (HiC-pro) are not provided  - we need to generate them from HiC-pro
WARNING: While bind mounting '/flask/scratch/briscoej/hamrude/atac_neural_plate_border/NF-hichip/run_fithichip/data:/flask/scratch/briscoej/hamrude/atac_neural_plate_border/NF-hichip/run_fithichip/data': destination is already in the mount point list

 ================ Parsing input configuration file =================
Content of ValidPairs is /flask/scratch/briscoej/thierya/atac_neural_plate_border/output/NF-hichip_alignment/hicpro/valid_pairs/NF_HiChip_r1.allValidPairs
Content of OutDir is /flask/scratch/briscoej/hamrude/atac_neural_plate_border/output/NF-hichip/NF_FitHiChiP_r1
Content of ChrSizeFile is /flask/scratch/briscoej/hamrude/atac_neural_plate_border/NF-hichip/run_fithichip/data/chrom_galgal6.sizes
Content of PeakFile is /flask/scratch/briscoej/hamrude/atac_neural_plate_border/output/NF-cutandrun-H3k27ac//03_peak_calling/05_consensus_peaks/h3k27ac.consensus.peaks.awk.bed
Content of CircularGenome is 0
Content of IntType is 3
Content of BINSIZE is 5000
Content of LowDistThr is 20000
Content of UppDistThr is 2000000
Content of QVALUE is 0.01
Content of UseP2PBackgrnd is 0
Content of BiasType is 1
Content of MergeInt is 1
Content of PREFIX is FitHiChIP
Content of OverWrite is 1

 ================ Verifying input configuration parameters =================
***** Specified output directory : /flask/scratch/briscoej/hamrude/atac_neural_plate_border/output/NF-hichip/NF_FitHiChiP_r1
HiC-pro is installed in the system
Installed HiC-pro version: 2.11.1
*** Valid python version is detected - installed version: 2.7.16
*** Python library gzip is installed
*** Python module OptionParser (from the package optparse) is installed
*** Python package networkx is installed
*** Found MACS2 package (for peak calling) installed in the system -  2.1.2
*** Valid R version is detected - installed R version: 3.6.1
*** Valid samtools version is detected - installed version: 1.9
*** bgzip utility is installed in the system
*** tabix utility is installed in the system
*** Valid bedtools version is detected - installed version: 2.28.0

 ================ Changing relative pathnames of the input files to their absolute path names =================
/flask/scratch/briscoej/hamrude/atac_neural_plate_border/NF-hichip/run_fithichip/data
/FitHiChIP/FitHiChIP_HiCPro.sh: line 841: [[: 0.01: syntax error: invalid arithmetic operator (error token is ".01")
/FitHiChIP/FitHiChIP_HiCPro.sh: line 841: [[: 0.01: syntax error: invalid arithmetic operator (error token is ".01")
Executable of python (2): /root/miniconda2/bin//python
Executable of R : /usr/bin/Rscript

 ================ Processing HiC-pro contact matrices =================

 *** MatrixBuildExec: /HiC-Pro-2.11.1/scripts/build_matrix
*** Computing HiC-pro matrices from the input valid pairs file
***** HiC-pro input valid pairs file in simple text format

 ================ Creating input interactions =================
Number of locus pairs with nonzero contact count (without any distance thresholding): 37450250

 ================ Limiting input interactions to the specified distance ranges 20000 to 2000000 =================
Number of locus pairs with nonzero contact count (after distance thresholding): 17533036

 ================ Generating coverage statistics for individual bins =================
Loading required package: stats4
Loading required package: BiocGenerics
Loading required package: parallel

Attaching package: ‘BiocGenerics’

The following objects are masked from ‘package:parallel’:

    clusterApply, clusterApplyLB, clusterCall, clusterEvalQ,
    clusterExport, clusterMap, parApply, parCapply, parLapply,
    parLapplyLB, parRapply, parSapply, parSapplyLB

The following objects are masked from ‘package:stats’:

    IQR, mad, sd, var, xtabs

The following objects are masked from ‘package:base’:

    anyDuplicated, append, as.data.frame, basename, cbind, colnames,
    dirname, do.call, duplicated, eval, evalq, Filter, Find, get, grep,
    grepl, intersect, is.unsorted, lapply, Map, mapply, match, mget,
    order, paste, pmax, pmax.int, pmin, pmin.int, Position, rank,
    rbind, Reduce, rownames, sapply, setdiff, sort, table, tapply,
    union, unique, unsplit, which, which.max, which.min

Loading required package: S4Vectors

Attaching package: ‘S4Vectors’

The following object is masked from ‘package:base’:

    expand.grid

Loading required package: IRanges
Loading required package: GenomeInfoDb
======== Computed initial coverage information for individual genomic bins

 ================ Merging coverage with bias statistics =================
======== Appended coverage bias information for individual genomic bins

 ================ Merging coverage + bias with mappability, GC content, and number of cut sites - creating all feature file =================

 ================ Generating interactions + features =================
Input interaction file: /flask/scratch/briscoej/hamrude/atac_neural_plate_border/output/NF-hichip/NF_FitHiChiP_r1/HiCPro_Matrix_BinSize5000/L_20000_U2000000/FitHiChIP.cis.interactions.DistThr.bed 
Output file: /flask/scratch/briscoej/hamrude/atac_neural_plate_border/output/NF-hichip/NF_FitHiChiP_r1/FitHiChIP_ALL2ALL_b5000_L20000_U2000000/Coverage_Bias/Interactions.bed 
File storing the normalization features for individual genomic intervals: /flask/scratch/briscoej/hamrude/atac_neural_plate_border/output/NF-hichip/NF_FitHiChiP_r1/NormFeatures/Coverage_Bias/FitHiChIP.AllBin_CompleteFeat.bed 

 Output directory: /flask/scratch/briscoej/hamrude/atac_neural_plate_border/output/NF-hichip/NF_FitHiChiP_r1/FitHiChIP_ALL2ALL_b5000_L20000_U2000000/Coverage_Bias 
Contact count col: 7
Total number of columns for the complete feature interactions: 19
Specified IntType: 3
Derived IntLow: 3
Derived IntHigh: 3


 **** Start of while Loop ----- current interaction type: 3  ****** 



 ============ Calling significant interactions =============== 

Created sorted genomic distance based interaction file
============== ************* Current directory for FitHiChIP significant interaction calling: /flask/scratch/briscoej/hamrude/atac_neural_plate_border/output/NF-hichip/NF_FitHiChiP_r1/FitHiChIP_Peak2ALL_b5000_L20000_U2000000/P2PBckgr_0/Coverage_Bias/FitHiC_BiasCorr

 Peak to peak background usage for spline fit: 0 
 Number of cores in the system: 48 
 Total Number of interactions: 8111055 
 ****** Total number of training interactions: 8111055 ********
 ****** Number of contacts per bin (allowed for equal occupancy binning): 103229 ******** 
Error in smooth.spline(avg_int_dist, prior_contact_prob, cv = TRUE) : 
  missing or infinite values in inputs are not allowed
Execution halted
******** FINISHED calling significant interactions
ERROR !!!!!!!! FitHiChIP could not compute the statistical significance of input interactions
Check the input parameters, or check if the number of input nonzero contact locus pairs are too few !!!
If you are using peak to peak background only (UseP2PBackgrnd=1 or stringent background), check the number of nonzero peak-to-peak locus pairs (peak-to-peak) !!! In such a case, you should go for loose background (UseP2PBackgrnd=0) for modeling interaction significance!!! Specially, if the sequencing depth of your data is very low....
awk: cannot open /flask/scratch/briscoej/hamrude/atac_neural_plate_border/output/NF-hichip/NF_FitHiChiP_r1/FitHiChIP_Peak2ALL_b5000_L20000_U2000000/P2PBckgr_0/Coverage_Bias/FitHiC_BiasCorr/FitHiChIP.interactions_FitHiC.bed (No such file or directory)
----- Extracted significant interactions ---- FDR threshold lower than: 0.01
SORRY !!!!!!!! FitHiChIP could not find any statistically significant interactions
Check the input parameters, or check if the number of input nonzero contact locus pairs are too few !!!
If you are using peak to peak background only (UseP2PBackgrnd=1 or stringent background), check the number of nonzero peak-to-peak locus pairs (peak-to-peak) !!! In such a case, you should go for loose background (UseP2PBackgrnd=0) for modeling interaction significance!!! Specially, if the sequencing depth of your data is very low....
There is no significant interaction - so no WashU specific session file is created !!
Updated CurrIntType: 4

 **** Now summarizing FitHiChIP results in the HTML file *** 

***** There are one or more errors / notifications (such as zero interaction count) associated with this execution !!!
***** Please check the console to know about them !!

Many thanks for your help,
Eva

[ay-lab]

Hi @evaham1
Thanks for your query. This is strange. Could you please email me ([email protected]) the validpairs, ChIP-seq peaks (only chromosome 1 is sufficient) and the chromosome size file? I'll debug this issue and let you know soon.