Problem: Estimating Optimal Numbers of Clusters #39
Comments
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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 |
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Hello all, I am getting the exact same issue as Maria. Here is my full log:
and this is where the error occurs:
My run command looks like this:
and my data source looks like this:
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. |
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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.
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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! |
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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!
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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)
Thank you for your help!
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