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https://rnbeads.org/
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Problem: Estimating Optimal Numbers of Clusters #39

Open mariacastillo982 opened 1 year ago

mariacastillo982 commented 1 year ago

Hello, I want to report that when I'm running my analysis, I'm getting the following error:

Error in dimnames(x) <- dn : length of 'dimnames' [2] not equal to array extent In addition: There were 50 or more warnings (use warnings() to see the first 50)

warnings() Warning messages: 1: The dot-dot notation (..count..) was deprecated in ggplot2 3.4.0. ℹ Please use after_stat(count) instead. ℹ The deprecated feature was likely used in the RnBeads package. Please report the issue to the authors. This warning is displayed once every 8 hours. Call lifecycle::last_lifecycle_warnings() to see where this warning was generated. 2: Removed 5370 rows containing non-finite values (stat_ydensity()). 3: Removed 1743 rows containing non-finite values (stat_density()). 4: Removed 1743 rows containing non-finite values (stat_density()). 5: Removed 627 rows containing non-finite values (stat_density()). 6: Removed 627 rows containing non-finite values (stat_density()). 7: In cor(t(x), use = "pairwise.complete.obs") : the standard deviation is zero 8: In cor(t(x), use = "pairwise.complete.obs") : the standard deviation is zero 9: In cor(t(x), use = "pairwise.complete.obs") : the standard deviation is zero 10: In cor(t(x), use = "pairwise.complete.obs") : the standard deviation is zero ... 50: In cor(t(x), use = "pairwise.complete.obs") : the standard deviation is zero

Thank you for your help!

schmic05 commented 1 year ago

Hi Maria,

Thanks for using RnBeads. From the message above, I cannot really tell where the issue arises from. Can you share the full analysis.log with us?

Thanks,

Michael

jcdaneshmand commented 8 months ago

Hello all, I am getting the exact same issue as Maria. Here is my full log:

2023-11-06 19:39:19 1.9 STATUS STARTED RnBeads Pipeline 2023-11-06 19:39:20 1.9 INFO Initialized report index and saved to index.html 2023-11-06 19:39:20 1.9 STATUS STARTED Loading Data 2023-11-06 19:39:20 1.9 INFO Number of cores: 1 2023-11-06 19:39:20 1.9 INFO Loading data of type "bs.bed.dir" 2023-11-06 19:39:20 1.9 STATUS STARTED Performing loading test 2023-11-06 19:39:20 1.9 INFO The first 10000 rows will be read from each data file 2023-11-06 19:39:20 1.9 INFO No column with file names specified: will try to find one 2023-11-06 19:39:20 1.9 STATUS STARTED Loading Data From BED Files 2023-11-06 19:39:21 2.0 STATUS STARTED Automatically parsing the provided sample annotation file 2023-11-06 19:39:21 2.0 STATUS Potential file names found in column 2 of the supplied annotation table 2023-11-06 19:39:21 2.0 STATUS COMPLETED Automatically parsing the provided sample annotation file 2023-11-06 19:39:21 2.0 INFO Reading BED file: /mnt/e/Repositories/Jonah/SFT_nextflow/methyl_results/bismark/methylation_calls/methylation_coverage/gunzipped/10303-AM-0001_S1_L005_1_val_1_bismark_bt2_pe.deduplicated.bismark.cov 2023-11-06 19:39:21 2.0 INFO Reading BED file: /mnt/e/Repositories/Jonah/SFT_nextflow/methyl_results/bismark/methylation_calls/methylation_coverage/gunzipped/10303-AM-0048_S1_L005_1_val_1_bismark_bt2_pe.deduplicated.bismark.cov 2023-11-06 19:39:22 2.0 INFO Reading BED file: /mnt/e/Repositories/Jonah/SFT_nextflow/methyl_results/bismark/methylation_calls/methylation_coverage/gunzipped/10303-AM-0125_S1_L005_1_val_1_bismark_bt2_pe.deduplicated.bismark.cov 2023-11-06 19:39:22 2.0 STATUS Read 3 BED files 2023-11-06 19:39:22 2.0 STATUS Matched chromosomes and strands to annotation 2023-11-06 19:39:22 2.0 STATUS Checked for the presence of sites and coverage 2023-11-06 19:39:22 2.0 STATUS Initialized meth/covg matrices 2023-11-06 19:39:23 2.0 STATUS Combined a data matrix with 14144 sites and 3 samples 2023-11-06 19:39:23 2.0 STATUS Processed all BED files 2023-11-06 19:39:23 2.0 STATUS STARTED Creating RnBiseqSet object 2023-11-06 19:39:23 2.0 INFO Inferring strand information from annotation enabled 2023-11-06 19:39:56 4.9 STATUS Matched 14111 of 14144 methylation sites to the annotation 2023-11-06 19:39:56 4.9 STATUS Checking site coverage 2023-11-06 19:39:56 4.9 STATUS Creating methylation matrix 2023-11-06 19:39:56 4.9 STATUS Creating coverage matrix 2023-11-06 19:39:56 4.9 STATUS Creating object 2023-11-06 19:39:56 4.9 STATUS Summarizing strand methylation 2023-11-06 19:39:59 5.0 STATUS Summarizing tiling methylation 2023-11-06 19:39:59 5.0 STATUS Summarizing genes methylation 2023-11-06 19:39:59 5.0 STATUS Summarizing promoters methylation 2023-11-06 19:39:59 5.0 STATUS Summarizing cpgislands methylation 2023-11-06 19:39:59 5.0 STATUS COMPLETED Creating RnBiseqSet object 2023-11-06 19:39:59 5.0 STATUS COMPLETED Loading Data From BED Files 2023-11-06 19:39:59 5.0 STATUS STARTED Checking the loaded object 2023-11-06 19:39:59 5.0 INFO Checking the supplied RnBiseqSet object 2023-11-06 19:39:59 5.0 INFO The object contains information for 8216 methylation sites 2023-11-06 19:39:59 5.0 INFO The object contains information for 3 samples 2023-11-06 19:39:59 5.0 INFO The object contains 6099 missing methylation values 2023-11-06 19:39:59 5.0 INFO Methylation values are within the expected range 2023-11-06 19:39:59 5.0 INFO The object contains coverage information 2023-11-06 19:39:59 5.0 INFO Coverage values are within the expected range 2023-11-06 19:39:59 5.0 INFO The object loaded during the loading test is valid 2023-11-06 19:39:59 5.0 STATUS COMPLETED Checking the loaded object 2023-11-06 19:39:59 5.0 STATUS COMPLETED Performing loading test 2023-11-06 19:39:59 5.0 INFO No column with file names specified: will try to find one 2023-11-06 19:39:59 5.0 STATUS STARTED Loading Data From BED Files 2023-11-06 19:39:59 5.0 STATUS STARTED Automatically parsing the provided sample annotation file 2023-11-06 19:39:59 5.0 STATUS Potential file names found in column 2 of the supplied annotation table 2023-11-06 19:39:59 5.0 STATUS COMPLETED Automatically parsing the provided sample annotation file 2023-11-06 19:39:59 5.0 INFO Reading BED file: /mnt/e/Repositories/Jonah/SFT_nextflow/methyl_results/bismark/methylation_calls/methylation_coverage/gunzipped/10303-AM-0001_S1_L005_1_val_1_bismark_bt2_pe.deduplicated.bismark.cov 2023-11-06 19:40:24 5.8 INFO Reading BED file: /mnt/e/Repositories/Jonah/SFT_nextflow/methyl_results/bismark/methylation_calls/methylation_coverage/gunzipped/10303-AM-0048_S1_L005_1_val_1_bismark_bt2_pe.deduplicated.bismark.cov 2023-11-06 19:40:43 5.3 INFO Reading BED file: /mnt/e/Repositories/Jonah/SFT_nextflow/methyl_results/bismark/methylation_calls/methylation_coverage/gunzipped/10303-AM-0125_S1_L005_1_val_1_bismark_bt2_pe.deduplicated.bismark.cov 2023-11-06 19:41:01 5.3 STATUS Read 3 BED files 2023-11-06 19:41:09 5.9 STATUS Matched chromosomes and strands to annotation 2023-11-06 19:41:09 5.9 STATUS Checked for the presence of sites and coverage 2023-11-06 19:41:10 6.1 STATUS Initialized meth/covg matrices 2023-11-06 19:41:16 5.7 STATUS Combined a data matrix with 10667924 sites and 3 samples 2023-11-06 19:41:16 5.7 STATUS Processed all BED files 2023-11-06 19:41:16 5.7 STATUS STARTED Creating RnBiseqSet object 2023-11-06 19:41:16 6.1 INFO Removed 26147 sites with unknown chromosomes 2023-11-06 19:41:16 5.9 INFO Inferring strand information from annotation enabled 2023-11-06 19:41:49 6.9 STATUS Matched 10595713 of 10667924 methylation sites to the annotation 2023-11-06 19:41:49 6.9 STATUS Checking site coverage 2023-11-06 19:41:52 6.8 STATUS Creating methylation matrix 2023-11-06 19:41:55 6.9 STATUS Creating coverage matrix 2023-11-06 19:41:58 7.1 STATUS Creating object 2023-11-06 19:41:59 6.5 STATUS Summarizing strand methylation 2023-11-06 19:43:37 7.1 STATUS Summarizing tiling methylation 2023-11-06 19:43:50 8.0 STATUS Summarizing genes methylation 2023-11-06 19:43:56 8.2 STATUS Summarizing promoters methylation 2023-11-06 19:44:01 8.2 STATUS Summarizing cpgislands methylation 2023-11-06 19:44:04 8.0 STATUS COMPLETED Creating RnBiseqSet object 2023-11-06 19:44:04 8.0 STATUS COMPLETED Loading Data From BED Files 2023-11-06 19:44:04 8.0 STATUS Loaded data from /mnt/e/Repositories/Jonah/SFT_nextflow/methyl_results/bismark/methylation_calls/methylation_coverage/gunzipped 2023-11-06 19:44:19 9.0 STATUS Predicted sex for the loaded samples 2023-11-06 19:44:19 9.0 STATUS Added data loading section to the report 2023-11-06 19:44:19 9.0 STATUS Loaded 3 samples and 6292277 sites 2023-11-06 19:44:19 9.0 INFO Output object is of type RnBiseqSet 2023-11-06 19:44:19 9.0 STATUS COMPLETED Loading Data 2023-11-06 19:44:25 9.0 INFO Initialized report index and saved to index.html 2023-11-06 19:44:25 9.0 STATUS STARTED Quality Control 2023-11-06 19:44:25 9.0 INFO Number of cores: 1 2023-11-06 19:44:25 9.0 STATUS STARTED Preparing Quality Control Information 2023-11-06 19:44:25 9.0 STATUS COMPLETED Preparing Quality Control Information 2023-11-06 19:44:25 9.0 STATUS STARTED Quality Control Section 2023-11-06 19:44:41 8.5 STATUS Added sequencing coverage histograms 2023-11-06 19:45:09 8.6 STATUS Added sample coverage section 2023-11-06 19:45:29 9.9 STATUS Added sequencing coverage violin plots 2023-11-06 19:45:32 9.7 STATUS COMPLETED Quality Control Section 2023-11-06 19:45:47 9.5 STATUS COMPLETED Quality Control 2023-11-06 19:45:48 9.5 INFO Initialized report index and saved to index.html 2023-11-06 19:45:48 9.5 STATUS STARTED Preprocessing 2023-11-06 19:45:48 9.5 INFO Number of cores: 1 2023-11-06 19:45:48 9.5 WARNING filtering.greedycut disabled for non-array datasets. 2023-11-06 19:45:48 9.5 STATUS STARTED Filtering Procedures 2023-11-06 19:45:53 9.9 STATUS STARTED Removal of SNP-enriched Sites 2023-11-06 19:45:53 9.9 STATUS Removed 46155 sites using SNP criterion "any" 2023-11-06 19:45:53 9.9 STATUS Saved removed sites to /mnt/e/Repositories/Jonah/SFT_MethylAnalysis/reports/preprocessing_data/removed_sites_snp.csv 2023-11-06 19:45:53 9.9 STATUS Added a corresponding section to the report 2023-11-06 19:45:53 9.9 STATUS COMPLETED Removal of SNP-enriched Sites 2023-11-06 19:45:53 9.9 STATUS STARTED Removal of Cross-reactive Probes 2023-11-06 19:45:53 9.9 STATUS Added a corresponding section to the report 2023-11-06 19:45:53 9.9 STATUS COMPLETED Removal of Cross-reactive Probes 2023-11-06 19:45:53 9.9 STATUS STARTED Removal of High Coverage (Outlier) Sites 2023-11-06 19:45:54 8.1 STATUS Removed 0 high coverage outlier sites 2023-11-06 19:45:54 8.1 STATUS Saved removed sites to /mnt/e/Repositories/Jonah/SFT_MethylAnalysis/reports/preprocessing_data/removed_sites_high_coverage.csv 2023-11-06 19:45:54 8.1 STATUS Added a corresponding section to the report 2023-11-06 19:45:54 8.1 STATUS COMPLETED Removal of High Coverage (Outlier) Sites 2023-11-06 19:45:54 8.1 STATUS STARTED Replacing Low Coverage Sites by NA 2023-11-06 19:45:55 9.5 STATUS Masked 6946071 site(s) based on coverage threshold 5 2023-11-06 19:45:56 9.5 STATUS Saved numbers of masked sites per sample to /mnt/e/Repositories/Jonah/SFT_MethylAnalysis/reports/preprocessing_data/masked_sites_coverage.csv 2023-11-06 19:45:56 9.5 STATUS Added a corresponding section to the report 2023-11-06 19:45:56 9.5 STATUS COMPLETED Replacing Low Coverage Sites by NA 2023-11-06 19:45:56 9.5 STATUS STARTED Removal of Sites on Sex Chromosomes 2023-11-06 19:45:56 9.6 STATUS Removed 160956 site(s) on sex chromosomes 2023-11-06 19:45:56 9.6 STATUS Saved removed sites to /mnt/e/Repositories/Jonah/SFT_MethylAnalysis/reports/preprocessing_data/removed_sites_sex.csv 2023-11-06 19:45:56 9.6 STATUS Added a corresponding section to the report 2023-11-06 19:45:56 9.6 STATUS COMPLETED Removal of Sites on Sex Chromosomes 2023-11-06 19:45:56 9.6 STATUS STARTED Missing Value Removal 2023-11-06 19:45:56 9.6 STATUS Using a sample quantile threshold of 0.5 2023-11-06 19:45:58 8.1 STATUS Removed 3395595 site(s) with too many missing values 2023-11-06 19:46:05 8.1 STATUS Saved removed sites to /mnt/e/Repositories/Jonah/SFT_MethylAnalysis/reports/preprocessing_data/removed_sites_na.csv 2023-11-06 19:46:08 9.4 STATUS Added a corresponding section to the report 2023-11-06 19:46:08 9.4 STATUS COMPLETED Missing Value Removal 2023-11-06 19:46:09 9.4 STATUS Retained 3 samples and 2689571 sites 2023-11-06 19:46:09 9.4 STATUS COMPLETED Filtering Procedures 2023-11-06 19:46:09 9.4 STATUS STARTED Summary of Filtering Procedures 2023-11-06 19:46:10 9.1 STATUS Created summary table of removed sites, samples and unreliable measurements 2023-11-06 19:46:10 9.1 STATUS Added summary table of removed and retained items 2023-11-06 19:46:11 9.1 INFO Subsampling 2000000 sites for plotting density distributions 2023-11-06 19:46:11 9.1 STATUS Constructed sequences of removed and retained methylation values 2023-11-06 19:46:14 9.1 STATUS Added comparison between removed and retained beta values 2023-11-06 19:46:14 9.1 STATUS COMPLETED Summary of Filtering Procedures 2023-11-06 19:46:14 9.1 STATUS STARTED Manipulating the object 2023-11-06 19:46:15 9.2 STATUS Updated NA masking 2023-11-06 19:46:29 9.3 STATUS Removed 3602706 sites (probes) 2023-11-06 19:46:29 9.3 INFO Retained 2689571 sites and 3 samples 2023-11-06 19:46:29 9.3 STATUS COMPLETED Manipulating the object 2023-11-06 19:46:29 9.3 INFO Imputation was skipped, data set may still contain missing methylation values 2023-11-06 19:46:29 9.3 STATUS COMPLETED Preprocessing 2023-11-06 19:46:32 9.3 INFO Initialized report index and saved to index.html 2023-11-06 19:46:32 9.3 STATUS STARTED Exploratory Analysis 2023-11-06 19:46:32 9.3 INFO Number of cores: 1 2023-11-06 19:46:33 9.3 STATUS Designed color mappings for probe type and CGI status 2023-11-06 19:47:20 9.0 STATUS STARTED Dimension Reduction Techniques 2023-11-06 19:47:20 9.0 WARNING Skipped due to too few samples 2023-11-06 19:47:20 9.0 STATUS COMPLETED Dimension Reduction Techniques 2023-11-06 19:47:24 9.5 STATUS STARTED Methylation Value Distributions - Sample Groups 2023-11-06 19:47:24 9.5 INFO processing beta_density_samples_1_1 2023-11-06 19:47:24 9.5 INFO Density estimation ( all samples--sites ): Groupwise retained observations after missing value removal: all:7062610/8068713 2023-11-06 19:47:24 9.5 INFO Density estimation ( all samples--sites ): Groupwise retained observations after subsampling: all:1000000/7062610 2023-11-06 19:47:26 9.5 INFO processing beta_density_samples_1_2 2023-11-06 19:47:26 9.5 INFO Density estimation ( all samples--tiling ): Groupwise retained observations after missing value removal: all:316269/348171 2023-11-06 19:47:26 9.5 INFO processing beta_density_samples_1_3 2023-11-06 19:47:26 9.5 INFO Density estimation ( all samples--genes ): Groupwise retained observations after missing value removal: all:87234/90342 2023-11-06 19:47:27 9.5 INFO processing beta_density_samples_1_4 2023-11-06 19:47:27 9.5 INFO Density estimation ( all samples--promoters ): Groupwise retained observations after missing value removal: all:95853/100704 2023-11-06 19:47:28 9.5 INFO processing beta_density_samples_1_5 2023-11-06 19:47:28 9.5 INFO Density estimation ( all samples--cpgislands ): Groupwise retained observations after missing value removal: all:76014/76956 2023-11-06 19:47:28 9.5 STATUS COMPLETED Methylation Value Distributions - Sample Groups 2023-11-06 19:47:28 9.5 STATUS STARTED Methylation Value Distributions - Site Categories 2023-11-06 19:47:28 9.5 INFO Density estimation ( CGI Relation--all samples ): Groupwise retained observations after missing value removal: Open Sea:2653048/3173163; Shelf:97996/116853; Shore:136662/163152; Island:4174904/4615545 2023-11-06 19:47:29 9.5 INFO Density estimation ( CGI Relation--all samples ): Groupwise retained observations after subsampling: Open Sea:1502588/2653048; Shelf:55501/97996; Shore:77400/136662; Island:2364511/4174904 2023-11-06 19:47:34 9.6 STATUS COMPLETED Methylation Value Distributions - Site Categories 2023-11-06 19:47:34 9.6 STATUS STARTED Sample Clustering 2023-11-06 19:47:34 9.6 STATUS STARTED Agglomerative Hierarchical Clustering 2023-11-06 19:47:34 9.6 STATUS Performed clustering on sites using correlation as a distance metric 2023-11-06 19:47:34 9.6 STATUS Performed clustering on sites using manhattan as a distance metric 2023-11-06 19:47:35 9.6 STATUS Performed clustering on sites using euclidean as a distance metric 2023-11-06 19:47:35 9.6 STATUS Performed clustering on tiling using correlation as a distance metric 2023-11-06 19:47:35 9.6 STATUS Performed clustering on tiling using manhattan as a distance metric 2023-11-06 19:47:35 9.6 STATUS Performed clustering on tiling using euclidean as a distance metric 2023-11-06 19:47:35 9.6 STATUS Performed clustering on genes using correlation as a distance metric 2023-11-06 19:47:35 9.6 STATUS Performed clustering on genes using manhattan as a distance metric 2023-11-06 19:47:35 9.6 STATUS Performed clustering on genes using euclidean as a distance metric 2023-11-06 19:47:35 9.6 STATUS Performed clustering on promoters using correlation as a distance metric 2023-11-06 19:47:35 9.6 STATUS Performed clustering on promoters using manhattan as a distance metric 2023-11-06 19:47:35 9.6 STATUS Performed clustering on promoters using euclidean as a distance metric 2023-11-06 19:47:35 9.6 STATUS Performed clustering on cpgislands using correlation as a distance metric 2023-11-06 19:47:35 9.6 STATUS Performed clustering on cpgislands using manhattan as a distance metric 2023-11-06 19:47:35 9.6 STATUS Performed clustering on cpgislands using euclidean as a distance metric 2023-11-06 19:47:35 9.6 STATUS COMPLETED Agglomerative Hierarchical Clustering 2023-11-06 19:47:35 9.6 STATUS STARTED Clustering Section 2023-11-06 19:47:35 9.6 STATUS STARTED Generating Heatmaps 2023-11-06 19:47:35 9.6 STATUS STARTED Region type: sites 2023-11-06 19:48:06 9.8 STATUS COMPLETED Region type: sites 2023-11-06 19:48:06 9.8 STATUS STARTED Region type: tiling 2023-11-06 19:48:35 9.8 STATUS COMPLETED Region type: tiling 2023-11-06 19:48:35 9.8 STATUS STARTED Region type: genes 2023-11-06 19:49:05 9.8 STATUS COMPLETED Region type: genes 2023-11-06 19:49:05 9.8 STATUS STARTED Region type: promoters 2023-11-06 19:49:35 9.8 STATUS COMPLETED Region type: promoters 2023-11-06 19:49:35 9.8 STATUS STARTED Region type: cpgislands 2023-11-06 19:50:05 9.2 STATUS COMPLETED Region type: cpgislands 2023-11-06 19:50:05 9.2 STATUS Created 135 heatmaps based on the clustering results 2023-11-06 19:50:05 9.2 STATUS COMPLETED Generating Heatmaps 2023-11-06 19:50:05 9.2 STATUS STARTED Adding Color Legends 2023-11-06 19:50:08 9.2 STATUS COMPLETED Adding Color Legends 2023-11-06 19:50:08 9.2 STATUS STARTED Estimating Optimal Numbers of Clusters

and this is where the error occurs:

Error in dimnames(x) <- dn : length of 'dimnames' [2] not equal to array extent In addition: There were 31 warnings (use warnings() to see them).

My run command looks like this:

rnb.run.analysis(dir.reports=report.dir, data.source=data_source, data.type="bs.bed.dir", initialize.reports = TRUE, save.rdata = TRUE)

and my data source looks like this:

str(data_source) List of 2 $ : chr "/mnt/e/Repositories/Jonah/SFT_nextflow/methyl_results/bismark/methylation_calls/methylation_coverage/gunzipped" $ :'data.frame': 3 obs. of 4 variables: ..$ SampleName : chr [1:3] "10303-AM-0001_S1_L005" "10303-AM-0048_S1_L005" "10303-AM-0125_S1_L005" ..$ FilePath : chr [1:3] "10303-AM-0001_S1_L005_1_val_1_bismark_bt2_pe.deduplicated.bismark.cov" "10303-AM-0048_S1_L005_1_val_1_bismark_bt2_pe.deduplicated.bismark.cov" "10303-AM-0125_S1_L005_1_val_1_bismark_bt2_pe.deduplicated.bismark.cov" ..$ group : chr [1:3] "SFT" "SFT" "MEN" ..$ group_Level2: chr [1:3] "iSFT" "eSFT" "MEN"

It runs for a while, and has some significant output in the reports folder, but the analysis does not finish. What am I doing wrong here? Thanks so much for any input.

mariacastillo982 commented 8 months ago

Hi, I think my issue was related to the number of samples I was using. I was analyzing only 3 samples, and there was a point where it was finding clusters, and I was getting that error. The way I could fix it was by increasing the number of samples I was using for the analysis and I stopped getting the error. I hope it helps you, otherwise the RnBeads is replying really fast.

Best wishes,

María de los Ángeles Gómez

Bioengineering

+966 (56) 650 5853

@. @.>

On Tue, Nov 7, 2023 at 4:07 AM Jonah Daneshmand @.***> wrote:

Hello all, I am getting the exact same issue as Maria. Here is my full log:

2023-11-06 19:39:19 1.9 STATUS STARTED RnBeads Pipeline 2023-11-06 19:39:20 1.9 INFO Initialized report index and saved to index.html 2023-11-06 19:39:20 1.9 STATUS STARTED Loading Data 2023-11-06 19:39:20 1.9 INFO Number of cores: 1 2023-11-06 19:39:20 1.9 INFO Loading data of type "bs.bed.dir" 2023-11-06 19:39:20 1.9 STATUS STARTED Performing loading test 2023-11-06 19:39:20 1.9 INFO The first 10000 rows will be read from each data file 2023-11-06 19:39:20 1.9 INFO No column with file names specified: will try to find one 2023-11-06 19:39:20 1.9 STATUS STARTED Loading Data From BED Files 2023-11-06 19:39:21 2.0 STATUS STARTED Automatically parsing the provided sample annotation file 2023-11-06 19:39:21 2.0 STATUS Potential file names found in column 2 of the supplied annotation table 2023-11-06 19:39:21 2.0 STATUS COMPLETED Automatically parsing the provided sample annotation file 2023-11-06 19:39:21 2.0 INFO Reading BED file: /mnt/e/Repositories/Jonah/SFT_nextflow/methyl_results/bismark/methylation_calls/methylation_coverage/gunzipped/10303-AM-0001_S1_L005_1_val_1_bismark_bt2_pe.deduplicated.bismark.cov 2023-11-06 19:39:21 2.0 INFO Reading BED file: /mnt/e/Repositories/Jonah/SFT_nextflow/methyl_results/bismark/methylation_calls/methylation_coverage/gunzipped/10303-AM-0048_S1_L005_1_val_1_bismark_bt2_pe.deduplicated.bismark.cov 2023-11-06 19:39:22 2.0 INFO Reading BED file: /mnt/e/Repositories/Jonah/SFT_nextflow/methyl_results/bismark/methylation_calls/methylation_coverage/gunzipped/10303-AM-0125_S1_L005_1_val_1_bismark_bt2_pe.deduplicated.bismark.cov 2023-11-06 19:39:22 2.0 STATUS Read 3 BED files 2023-11-06 19:39:22 2.0 STATUS Matched chromosomes and strands to annotation 2023-11-06 19:39:22 2.0 STATUS Checked for the presence of sites and coverage 2023-11-06 19:39:22 2.0 STATUS Initialized meth/covg matrices 2023-11-06 19:39:23 2.0 STATUS Combined a data matrix with 14144 sites and 3 samples 2023-11-06 19:39:23 2.0 STATUS Processed all BED files 2023-11-06 19:39:23 2.0 STATUS STARTED Creating RnBiseqSet object 2023-11-06 19:39:23 2.0 INFO Inferring strand information from annotation enabled 2023-11-06 19:39:56 4.9 STATUS Matched 14111 of 14144 methylation sites to the annotation 2023-11-06 19:39:56 4.9 STATUS Checking site coverage 2023-11-06 19:39:56 4.9 STATUS Creating methylation matrix 2023-11-06 19:39:56 4.9 STATUS Creating coverage matrix 2023-11-06 19:39:56 4.9 STATUS Creating object 2023-11-06 19:39:56 4.9 STATUS Summarizing strand methylation 2023-11-06 19:39:59 5.0 STATUS Summarizing tiling methylation 2023-11-06 19:39:59 5.0 STATUS Summarizing genes methylation 2023-11-06 19:39:59 5.0 STATUS Summarizing promoters methylation 2023-11-06 19:39:59 5.0 STATUS Summarizing cpgislands methylation 2023-11-06 19:39:59 5.0 STATUS COMPLETED Creating RnBiseqSet object 2023-11-06 19:39:59 5.0 STATUS COMPLETED Loading Data From BED Files 2023-11-06 19:39:59 5.0 STATUS STARTED Checking the loaded object 2023-11-06 19:39:59 5.0 INFO Checking the supplied RnBiseqSet object 2023-11-06 19:39:59 5.0 INFO The object contains information for 8216 methylation sites 2023-11-06 19:39:59 5.0 INFO The object contains information for 3 samples 2023-11-06 19:39:59 5.0 INFO The object contains 6099 missing methylation values 2023-11-06 19:39:59 5.0 INFO Methylation values are within the expected range 2023-11-06 19:39:59 5.0 INFO The object contains coverage information 2023-11-06 19:39:59 5.0 INFO Coverage values are within the expected range 2023-11-06 19:39:59 5.0 INFO The object loaded during the loading test is valid 2023-11-06 19:39:59 5.0 STATUS COMPLETED Checking the loaded object 2023-11-06 19:39:59 5.0 STATUS COMPLETED Performing loading test 2023-11-06 19:39:59 5.0 INFO No column with file names specified: will try to find one 2023-11-06 19:39:59 5.0 STATUS STARTED Loading Data From BED Files 2023-11-06 19:39:59 5.0 STATUS STARTED Automatically parsing the provided sample annotation file 2023-11-06 19:39:59 5.0 STATUS Potential file names found in column 2 of the supplied annotation table 2023-11-06 19:39:59 5.0 STATUS COMPLETED Automatically parsing the provided sample annotation file 2023-11-06 19:39:59 5.0 INFO Reading BED file: /mnt/e/Repositories/Jonah/SFT_nextflow/methyl_results/bismark/methylation_calls/methylation_coverage/gunzipped/10303-AM-0001_S1_L005_1_val_1_bismark_bt2_pe.deduplicated.bismark.cov 2023-11-06 19:40:24 5.8 INFO Reading BED file: /mnt/e/Repositories/Jonah/SFT_nextflow/methyl_results/bismark/methylation_calls/methylation_coverage/gunzipped/10303-AM-0048_S1_L005_1_val_1_bismark_bt2_pe.deduplicated.bismark.cov 2023-11-06 19:40:43 5.3 INFO Reading BED file: /mnt/e/Repositories/Jonah/SFT_nextflow/methyl_results/bismark/methylation_calls/methylation_coverage/gunzipped/10303-AM-0125_S1_L005_1_val_1_bismark_bt2_pe.deduplicated.bismark.cov 2023-11-06 19:41:01 5.3 STATUS Read 3 BED files 2023-11-06 19:41:09 5.9 STATUS Matched chromosomes and strands to annotation 2023-11-06 19:41:09 5.9 STATUS Checked for the presence of sites and coverage 2023-11-06 19:41:10 6.1 STATUS Initialized meth/covg matrices 2023-11-06 19:41:16 5.7 STATUS Combined a data matrix with 10667924 sites and 3 samples 2023-11-06 19:41:16 5.7 STATUS Processed all BED files 2023-11-06 19:41:16 5.7 STATUS STARTED Creating RnBiseqSet object 2023-11-06 19:41:16 6.1 INFO Removed 26147 sites with unknown chromosomes 2023-11-06 19:41:16 5.9 INFO Inferring strand information from annotation enabled 2023-11-06 19:41:49 6.9 STATUS Matched 10595713 of 10667924 methylation sites to the annotation 2023-11-06 19:41:49 6.9 STATUS Checking site coverage 2023-11-06 19:41:52 6.8 STATUS Creating methylation matrix 2023-11-06 19:41:55 6.9 STATUS Creating coverage matrix 2023-11-06 19:41:58 7.1 STATUS Creating object 2023-11-06 19:41:59 6.5 STATUS Summarizing strand methylation 2023-11-06 19:43:37 7.1 STATUS Summarizing tiling methylation 2023-11-06 19:43:50 8.0 STATUS Summarizing genes methylation 2023-11-06 19:43:56 8.2 STATUS Summarizing promoters methylation 2023-11-06 19:44:01 8.2 STATUS Summarizing cpgislands methylation 2023-11-06 19:44:04 8.0 STATUS COMPLETED Creating RnBiseqSet object 2023-11-06 19:44:04 8.0 STATUS COMPLETED Loading Data From BED Files 2023-11-06 19:44:04 8.0 STATUS Loaded data from /mnt/e/Repositories/Jonah/SFT_nextflow/methyl_results/bismark/methylation_calls/methylation_coverage/gunzipped 2023-11-06 19:44:19 9.0 STATUS Predicted sex for the loaded samples 2023-11-06 19:44:19 9.0 STATUS Added data loading section to the report 2023-11-06 19:44:19 9.0 STATUS Loaded 3 samples and 6292277 sites 2023-11-06 19:44:19 9.0 INFO Output object is of type RnBiseqSet 2023-11-06 19:44:19 9.0 STATUS COMPLETED Loading Data 2023-11-06 19:44:25 9.0 INFO Initialized report index and saved to index.html 2023-11-06 19:44:25 9.0 STATUS STARTED Quality Control 2023-11-06 19:44:25 9.0 INFO Number of cores: 1 2023-11-06 19:44:25 9.0 STATUS STARTED Preparing Quality Control Information 2023-11-06 19:44:25 9.0 STATUS COMPLETED Preparing Quality Control Information 2023-11-06 19:44:25 9.0 STATUS STARTED Quality Control Section 2023-11-06 19:44:41 8.5 STATUS Added sequencing coverage histograms 2023-11-06 19:45:09 8.6 STATUS Added sample coverage section 2023-11-06 19:45:29 9.9 STATUS Added sequencing coverage violin plots 2023-11-06 19:45:32 9.7 STATUS COMPLETED Quality Control Section 2023-11-06 19:45:47 9.5 STATUS COMPLETED Quality Control 2023-11-06 19:45:48 9.5 INFO Initialized report index and saved to index.html 2023-11-06 19:45:48 9.5 STATUS STARTED Preprocessing 2023-11-06 19:45:48 9.5 INFO Number of cores: 1 2023-11-06 19:45:48 9.5 WARNING filtering.greedycut disabled for non-array datasets. 2023-11-06 19:45:48 9.5 STATUS STARTED Filtering Procedures 2023-11-06 19:45:53 9.9 STATUS STARTED Removal of SNP-enriched Sites 2023-11-06 19:45:53 9.9 STATUS Removed 46155 sites using SNP criterion "any" 2023-11-06 19:45:53 9.9 STATUS Saved removed sites to /mnt/e/Repositories/Jonah/SFT_MethylAnalysis/reports/preprocessing_data/removed_sites_snp.csv 2023-11-06 19:45:53 9.9 STATUS Added a corresponding section to the report 2023-11-06 19:45:53 9.9 STATUS COMPLETED Removal of SNP-enriched Sites 2023-11-06 19:45:53 9.9 STATUS STARTED Removal of Cross-reactive Probes 2023-11-06 19:45:53 9.9 STATUS Added a corresponding section to the report 2023-11-06 19:45:53 9.9 STATUS COMPLETED Removal of Cross-reactive Probes 2023-11-06 19:45:53 9.9 STATUS STARTED Removal of High Coverage (Outlier) Sites 2023-11-06 19:45:54 8.1 STATUS Removed 0 high coverage outlier sites 2023-11-06 19:45:54 8.1 STATUS Saved removed sites to /mnt/e/Repositories/Jonah/SFT_MethylAnalysis/reports/preprocessing_data/removed_sites_high_coverage.csv 2023-11-06 19:45:54 8.1 STATUS Added a corresponding section to the report 2023-11-06 19:45:54 8.1 STATUS COMPLETED Removal of High Coverage (Outlier) Sites 2023-11-06 19:45:54 8.1 STATUS STARTED Replacing Low Coverage Sites by NA 2023-11-06 19:45:55 9.5 STATUS Masked 6946071 site(s) based on coverage threshold 5 2023-11-06 19:45:56 9.5 STATUS Saved numbers of masked sites per sample to /mnt/e/Repositories/Jonah/SFT_MethylAnalysis/reports/preprocessing_data/masked_sites_coverage.csv 2023-11-06 19:45:56 9.5 STATUS Added a corresponding section to the report 2023-11-06 19:45:56 9.5 STATUS COMPLETED Replacing Low Coverage Sites by NA 2023-11-06 19:45:56 9.5 STATUS STARTED Removal of Sites on Sex Chromosomes 2023-11-06 19:45:56 9.6 STATUS Removed 160956 site(s) on sex chromosomes 2023-11-06 19:45:56 9.6 STATUS Saved removed sites to /mnt/e/Repositories/Jonah/SFT_MethylAnalysis/reports/preprocessing_data/removed_sites_sex.csv 2023-11-06 19:45:56 9.6 STATUS Added a corresponding section to the report 2023-11-06 19:45:56 9.6 STATUS COMPLETED Removal of Sites on Sex Chromosomes 2023-11-06 19:45:56 9.6 STATUS STARTED Missing Value Removal 2023-11-06 19:45:56 9.6 STATUS Using a sample quantile threshold of 0.5 2023-11-06 19:45:58 8.1 STATUS Removed 3395595 site(s) with too many missing values 2023-11-06 19:46:05 8.1 STATUS Saved removed sites to /mnt/e/Repositories/Jonah/SFT_MethylAnalysis/reports/preprocessing_data/removed_sites_na.csv 2023-11-06 19:46:08 9.4 STATUS Added a corresponding section to the report 2023-11-06 19:46:08 9.4 STATUS COMPLETED Missing Value Removal 2023-11-06 19:46:09 9.4 STATUS Retained 3 samples and 2689571 sites 2023-11-06 19:46:09 9.4 STATUS COMPLETED Filtering Procedures 2023-11-06 19:46:09 9.4 STATUS STARTED Summary of Filtering Procedures 2023-11-06 19:46:10 9.1 STATUS Created summary table of removed sites, samples and unreliable measurements 2023-11-06 19:46:10 9.1 STATUS Added summary table of removed and retained items 2023-11-06 19:46:11 9.1 INFO Subsampling 2000000 sites for plotting density distributions 2023-11-06 19:46:11 9.1 STATUS Constructed sequences of removed and retained methylation values 2023-11-06 19:46:14 9.1 STATUS Added comparison between removed and retained beta values 2023-11-06 19:46:14 9.1 STATUS COMPLETED Summary of Filtering Procedures 2023-11-06 19:46:14 9.1 STATUS STARTED Manipulating the object 2023-11-06 19:46:15 9.2 STATUS Updated NA masking 2023-11-06 19:46:29 9.3 STATUS Removed 3602706 sites (probes) 2023-11-06 19:46:29 9.3 INFO Retained 2689571 sites and 3 samples 2023-11-06 19:46:29 9.3 STATUS COMPLETED Manipulating the object 2023-11-06 19:46:29 9.3 INFO Imputation was skipped, data set may still contain missing methylation values 2023-11-06 19:46:29 9.3 STATUS COMPLETED Preprocessing 2023-11-06 19:46:32 9.3 INFO Initialized report index and saved to index.html 2023-11-06 19:46:32 9.3 STATUS STARTED Exploratory Analysis 2023-11-06 19:46:32 9.3 INFO Number of cores: 1 2023-11-06 19:46:33 9.3 STATUS Designed color mappings for probe type and CGI status 2023-11-06 19:47:20 9.0 STATUS STARTED Dimension Reduction Techniques 2023-11-06 19:47:20 9.0 WARNING Skipped due to too few samples 2023-11-06 19:47:20 9.0 STATUS COMPLETED Dimension Reduction Techniques 2023-11-06 19:47:24 9.5 STATUS STARTED Methylation Value Distributions - Sample Groups 2023-11-06 19:47:24 9.5 INFO processing beta_density_samples_1_1 2023-11-06 19:47:24 9.5 INFO Density estimation ( all samples--sites ): Groupwise retained observations after missing value removal: all:7062610/8068713 2023-11-06 19:47:24 9.5 INFO Density estimation ( all samples--sites ): Groupwise retained observations after subsampling: all:1000000/7062610 2023-11-06 19:47:26 9.5 INFO processing beta_density_samples_1_2 2023-11-06 19:47:26 9.5 INFO Density estimation ( all samples--tiling ): Groupwise retained observations after missing value removal: all:316269/348171 2023-11-06 19:47:26 9.5 INFO processing beta_density_samples_1_3 2023-11-06 19:47:26 9.5 INFO Density estimation ( all samples--genes ): Groupwise retained observations after missing value removal: all:87234/90342 2023-11-06 19:47:27 9.5 INFO processing beta_density_samples_1_4 2023-11-06 19:47:27 9.5 INFO Density estimation ( all samples--promoters ): Groupwise retained observations after missing value removal: all:95853/100704 2023-11-06 19:47:28 9.5 INFO processing beta_density_samples_1_5 2023-11-06 19:47:28 9.5 INFO Density estimation ( all samples--cpgislands ): Groupwise retained observations after missing value removal: all:76014/76956 2023-11-06 19:47:28 9.5 STATUS COMPLETED Methylation Value Distributions - Sample Groups 2023-11-06 19:47:28 9.5 STATUS STARTED Methylation Value Distributions - Site Categories 2023-11-06 19:47:28 9.5 INFO Density estimation ( CGI Relation--all samples ): Groupwise retained observations after missing value removal: Open Sea:2653048/3173163; Shelf:97996/116853; Shore:136662/163152; Island:4174904/4615545 2023-11-06 19:47:29 9.5 INFO Density estimation ( CGI Relation--all samples ): Groupwise retained observations after subsampling: Open Sea:1502588/2653048; Shelf:55501/97996; Shore:77400/136662; Island:2364511/4174904 2023-11-06 19:47:34 9.6 STATUS COMPLETED Methylation Value Distributions - Site Categories 2023-11-06 19:47:34 9.6 STATUS STARTED Sample Clustering 2023-11-06 19:47:34 9.6 STATUS STARTED Agglomerative Hierarchical Clustering 2023-11-06 19:47:34 9.6 STATUS Performed clustering on sites using correlation as a distance metric 2023-11-06 19:47:34 9.6 STATUS Performed clustering on sites using manhattan as a distance metric 2023-11-06 19:47:35 9.6 STATUS Performed clustering on sites using euclidean as a distance metric 2023-11-06 19:47:35 9.6 STATUS Performed clustering on tiling using correlation as a distance metric 2023-11-06 19:47:35 9.6 STATUS Performed clustering on tiling using manhattan as a distance metric 2023-11-06 19:47:35 9.6 STATUS Performed clustering on tiling using euclidean as a distance metric 2023-11-06 19:47:35 9.6 STATUS Performed clustering on genes using correlation as a distance metric 2023-11-06 19:47:35 9.6 STATUS Performed clustering on genes using manhattan as a distance metric 2023-11-06 19:47:35 9.6 STATUS Performed clustering on genes using euclidean as a distance metric 2023-11-06 19:47:35 9.6 STATUS Performed clustering on promoters using correlation as a distance metric 2023-11-06 19:47:35 9.6 STATUS Performed clustering on promoters using manhattan as a distance metric 2023-11-06 19:47:35 9.6 STATUS Performed clustering on promoters using euclidean as a distance metric 2023-11-06 19:47:35 9.6 STATUS Performed clustering on cpgislands using correlation as a distance metric 2023-11-06 19:47:35 9.6 STATUS Performed clustering on cpgislands using manhattan as a distance metric 2023-11-06 19:47:35 9.6 STATUS Performed clustering on cpgislands using euclidean as a distance metric 2023-11-06 19:47:35 9.6 STATUS COMPLETED Agglomerative Hierarchical Clustering 2023-11-06 19:47:35 9.6 STATUS STARTED Clustering Section 2023-11-06 19:47:35 9.6 STATUS STARTED Generating Heatmaps 2023-11-06 19:47:35 9.6 STATUS STARTED Region type: sites 2023-11-06 19:48:06 9.8 STATUS COMPLETED Region type: sites 2023-11-06 19:48:06 9.8 STATUS STARTED Region type: tiling 2023-11-06 19:48:35 9.8 STATUS COMPLETED Region type: tiling 2023-11-06 19:48:35 9.8 STATUS STARTED Region type: genes 2023-11-06 19:49:05 9.8 STATUS COMPLETED Region type: genes 2023-11-06 19:49:05 9.8 STATUS STARTED Region type: promoters 2023-11-06 19:49:35 9.8 STATUS COMPLETED Region type: promoters 2023-11-06 19:49:35 9.8 STATUS STARTED Region type: cpgislands 2023-11-06 19:50:05 9.2 STATUS COMPLETED Region type: cpgislands 2023-11-06 19:50:05 9.2 STATUS Created 135 heatmaps based on the clustering results 2023-11-06 19:50:05 9.2 STATUS COMPLETED Generating Heatmaps 2023-11-06 19:50:05 9.2 STATUS STARTED Adding Color Legends 2023-11-06 19:50:08 9.2 STATUS COMPLETED Adding Color Legends 2023-11-06 19:50:08 9.2 STATUS STARTED Estimating Optimal Numbers of Clusters

and this is where the error occurs:

Error in dimnames(x) <- dn : length of 'dimnames' [2] not equal to array extent In addition: There were 31 warnings (use warnings() to see them).

My run command looks like this:

rnb.run.analysis(dir.reports=report.dir, data.source=data_source, data.type="bs.bed.dir", initialize.reports = TRUE, save.rdata = TRUE)

and my data source looks like this:

str(data_source) List of 2 $ : chr "/mnt/e/Repositories/Jonah/SFT_nextflow/methyl_results/bismark/methylation_calls/methylation_coverage/gunzipped" $ :'data.frame': 3 obs. of 4 variables: ..$ SampleName : chr [1:3] "10303-AM-0001_S1_L005" "10303-AM-0048_S1_L005" "10303-AM-0125_S1_L005" ..$ FilePath : chr [1:3] "10303-AM-0001_S1_L005_1_val_1_bismark_bt2_pe.deduplicated.bismark.cov" "10303-AM-0048_S1_L005_1_val_1_bismark_bt2_pe.deduplicated.bismark.cov" "10303-AM-0125_S1_L005_1_val_1_bismark_bt2_pe.deduplicated.bismark.cov" ..$ group : chr [1:3] "SFT" "SFT" "MEN" ..$ group_Level2: chr [1:3] "iSFT" "eSFT" "MEN"

It runs for a while, and has some significant output in the reports folder, but the analysis does not finish. What am I doing wrong here? Thanks so much for any input.

— Reply to this email directly, view it on GitHub https://urldefense.com/v3/__https://github.com/epigen/RnBeads/issues/39*issuecomment-1797110576__;Iw!!Nmw4Hv0!3ivn965GiAt9HTeE4whW9xYvVNiiuAZxCxwB-Oa27Yf4c6VQGxsfJkFW6RWJoIEuYuFqGSKTCQN9qyV098oLVRvnjHW2Pxp5$, or unsubscribe https://urldefense.com/v3/__https://github.com/notifications/unsubscribe-auth/AXRTMXUUR23UZ4Y5TOTXZODYDGCWXAVCNFSM6AAAAAAXMCKO7KVHI2DSMVQWIX3LMV43OSLTON2WKQ3PNVWWK3TUHMYTOOJXGEYTANJXGY__;!!Nmw4Hv0!3ivn965GiAt9HTeE4whW9xYvVNiiuAZxCxwB-Oa27Yf4c6VQGxsfJkFW6RWJoIEuYuFqGSKTCQN9qyV098oLVRvnjJvCuTEw$ . You are receiving this because you authored the thread.Message ID: @.***>

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jcdaneshmand commented 8 months ago

Thank you @mariacastillo982 ! That turned out to be my problem as well and makes total sense. I tested the pipeline with 9 samples instead and it worked. Cheers!

mariacastillo982 commented 8 months ago

Glad to hear it! Wish you the best in your analysis

On Sat, 11 Nov 2023 at 6:10 PM Jonah Daneshmand @.***> wrote:

Thank you @mariacastillo982 https://urldefense.com/v3/__https://github.com/mariacastillo982__;!!Nmw4Hv0!y6s8Ci8zgWcE3GRAOPeNvDb4rVPdDRrJiCCAIJG00YdAX52VXgNicjTDQcBXOVr838UaHW6GsSH5D66leOokWdvi-wIof5e4$ ! That turned out to be my problem as well and makes total sense. I tested the pipeline with 9 samples instead and it worked. Cheers!

— Reply to this email directly, view it on GitHub https://urldefense.com/v3/__https://github.com/epigen/RnBeads/issues/39*issuecomment-1806840355__;Iw!!Nmw4Hv0!y6s8Ci8zgWcE3GRAOPeNvDb4rVPdDRrJiCCAIJG00YdAX52VXgNicjTDQcBXOVr838UaHW6GsSH5D66leOokWdvi-4fhmC4Q$, or unsubscribe https://urldefense.com/v3/__https://github.com/notifications/unsubscribe-auth/AXRTMXVL3PDYAZED7X6PWV3YD6IORAVCNFSM6AAAAAAXMCKO7KVHI2DSMVQWIX3LMV43OSLTON2WKQ3PNVWWK3TUHMYTQMBWHA2DAMZVGU__;!!Nmw4Hv0!y6s8Ci8zgWcE3GRAOPeNvDb4rVPdDRrJiCCAIJG00YdAX52VXgNicjTDQcBXOVr838UaHW6GsSH5D66leOokWdvi-05Mo8TZ$ . You are receiving this because you were mentioned.Message ID: @.***>

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