Novakovic et al 2019 reanalysis
The methylation in ART meta-analysis group
2024-01-03
source("meth_functions.R")Obtaining array annotations
anno <- getAnnotation(IlluminaHumanMethylationEPICanno.ilm10b4.hg19)
myann <- data.frame(anno[,c("UCSC_RefGene_Name","Regulatory_Feature_Group","Islands_Name","Relation_to_Island")])
promoters <- grep("Prom",myann$Regulatory_Feature_Group)Introduction
This report is the reanalysis of first analysed DNA methylation data by [Novakovic et al (2019)] (https://doi.org/10.1038/s41467-019-11929-9)
In this study, DNA methylation status was generated for 149 neonatal (84♀ 65♂) and 158 adult (87♀ 71♂) ART-conceived individuals and for 58 neonatal (37♀, 21♂) and 75 adult (51♀, 24♂) non-ART conceived individuals.
WORKING_DIR="GSE131433"
ARRAY_DATA="GSE131433_RAW.tar"
DEST=paste(WORKING_DIR,"/",ARRAY_DATA,sep="")
IDAT="GSE131433/IDAT"
options(timeout=1000000)
if(!dir.exists(WORKING_DIR)){
dir.create(WORKING_DIR)
}
if(!file.exists(DEST)){
download.file("https://www.ncbi.nlm.nih.gov/geo/download/?acc=GSE131433&format=file",
# download.file("ftp://ftp.ncbi.nlm.nih.gov/geo/series/GSE131nnn/GSE131433/suppl/GSE131433_RAW.tar",
destfile=DEST)
untar(exdir = "IDAT", tarfile = DEST)
}
SERIES_MATRIX=paste(WORKING_DIR,"/","GSE131433_series_matrix.txt.gz",sep="")
if(!file.exists(SERIES_MATRIX)){
download.file("ftp://ftp.ncbi.nlm.nih.gov/geo/series/GSE131nnn/GSE131433/matrix/GSE131433_series_matrix.txt.gz",
destfile=SERIES_MATRIX)
}
gse <- getGEO(filename=SERIES_MATRIX)
baseDir <- "."
sample_metadata <- pData(phenoData(gse))
targets <- sample_metadata
targets <- subset(targets,`time of collection:ch1`=="birth")
targets$Basename <- head( gsub("_Grn.idat.gz","",list.files("IDAT",full.names=TRUE,pattern="Grn")) , 207 )
head(targets)## title geo_accession status
## GSM3780106 neonate_ART_Donor_1 GSM3780106 Public on Jul 29 2019
## GSM3780107 neonate_ART_Donor_2 GSM3780107 Public on Jul 29 2019
## GSM3780108 neonate_ART_Donor_3 GSM3780108 Public on Jul 29 2019
## GSM3780109 neonate_ART_Donor_4 GSM3780109 Public on Jul 29 2019
## GSM3780110 neonate_ART_Donor_5 GSM3780110 Public on Jul 29 2019
## GSM3780111 neonate_ART_Donor_6 GSM3780111 Public on Jul 29 2019
## submission_date last_update_date type channel_count
## GSM3780106 May 17 2019 Mar 20 2023 genomic 1
## GSM3780107 May 17 2019 Mar 20 2023 genomic 1
## GSM3780108 May 17 2019 Mar 20 2023 genomic 1
## GSM3780109 May 17 2019 Mar 20 2023 genomic 1
## GSM3780110 May 17 2019 Mar 20 2023 genomic 1
## GSM3780111 May 17 2019 Mar 20 2023 genomic 1
## source_name_ch1 organism_ch1 characteristics_ch1
## GSM3780106 ART_Donor_1 Homo sapiens gender: F
## GSM3780107 ART_Donor_2 Homo sapiens gender: M
## GSM3780108 ART_Donor_3 Homo sapiens gender: F
## GSM3780109 ART_Donor_4 Homo sapiens gender: M
## GSM3780110 ART_Donor_5 Homo sapiens gender: F
## GSM3780111 ART_Donor_6 Homo sapiens gender: F
## characteristics_ch1.1 characteristics_ch1.2
## GSM3780106 sample type: Guthrie blood spot art status: ART
## GSM3780107 sample type: Guthrie blood spot art status: ART
## GSM3780108 sample type: Guthrie blood spot art status: ART
## GSM3780109 sample type: Guthrie blood spot art status: ART
## GSM3780110 sample type: Guthrie blood spot art status: ART
## GSM3780111 sample type: Guthrie blood spot art status: ART
## characteristics_ch1.3 characteristics_ch1.4 molecule_ch1
## GSM3780106 art subtype: Frozen embryo time of collection: birth genomic DNA
## GSM3780107 art subtype: NA time of collection: birth genomic DNA
## GSM3780108 art subtype: Fresh embryo time of collection: birth genomic DNA
## GSM3780109 art subtype: Frozen embryo time of collection: birth genomic DNA
## GSM3780110 art subtype: Fresh embryo time of collection: birth genomic DNA
## GSM3780111 art subtype: Fresh embryo time of collection: birth genomic DNA
## extract_protocol_ch1
## GSM3780106 DNA was extracted using the Zymo Research ZR DNA-Card Extraction Kit following manufacturer protocol
## GSM3780107 DNA was extracted using the Zymo Research ZR DNA-Card Extraction Kit following manufacturer protocol
## GSM3780108 DNA was extracted using the Zymo Research ZR DNA-Card Extraction Kit following manufacturer protocol
## GSM3780109 DNA was extracted using the Zymo Research ZR DNA-Card Extraction Kit following manufacturer protocol
## GSM3780110 DNA was extracted using the Zymo Research ZR DNA-Card Extraction Kit following manufacturer protocol
## GSM3780111 DNA was extracted using the Zymo Research ZR DNA-Card Extraction Kit following manufacturer protocol
## label_ch1 label_protocol_ch1 taxid_ch1
## GSM3780106 Cy5 and Cy3 Standard Illumina Protocol 9606
## GSM3780107 Cy5 and Cy3 Standard Illumina Protocol 9606
## GSM3780108 Cy5 and Cy3 Standard Illumina Protocol 9606
## GSM3780109 Cy5 and Cy3 Standard Illumina Protocol 9606
## GSM3780110 Cy5 and Cy3 Standard Illumina Protocol 9606
## GSM3780111 Cy5 and Cy3 Standard Illumina Protocol 9606
## hyb_protocol
## GSM3780106 bisulphite converted DNA was amplified, fragmented and hybridised to Illumina Infinium MethylationEPIC Beadchip using standard Illumina protocol
## GSM3780107 bisulphite converted DNA was amplified, fragmented and hybridised to Illumina Infinium MethylationEPIC Beadchip using standard Illumina protocol
## GSM3780108 bisulphite converted DNA was amplified, fragmented and hybridised to Illumina Infinium MethylationEPIC Beadchip using standard Illumina protocol
## GSM3780109 bisulphite converted DNA was amplified, fragmented and hybridised to Illumina Infinium MethylationEPIC Beadchip using standard Illumina protocol
## GSM3780110 bisulphite converted DNA was amplified, fragmented and hybridised to Illumina Infinium MethylationEPIC Beadchip using standard Illumina protocol
## GSM3780111 bisulphite converted DNA was amplified, fragmented and hybridised to Illumina Infinium MethylationEPIC Beadchip using standard Illumina protocol
## scan_protocol
## GSM3780106 Arrays were imaged using BeadArray Reader using standard recommended Illumina scanner setting
## GSM3780107 Arrays were imaged using BeadArray Reader using standard recommended Illumina scanner setting
## GSM3780108 Arrays were imaged using BeadArray Reader using standard recommended Illumina scanner setting
## GSM3780109 Arrays were imaged using BeadArray Reader using standard recommended Illumina scanner setting
## GSM3780110 Arrays were imaged using BeadArray Reader using standard recommended Illumina scanner setting
## GSM3780111 Arrays were imaged using BeadArray Reader using standard recommended Illumina scanner setting
## description
## GSM3780106 guthrie blood spot from ART conceived individual
## GSM3780107 guthrie blood spot from ART conceived individual
## GSM3780108 guthrie blood spot from ART conceived individual
## GSM3780109 guthrie blood spot from ART conceived individual
## GSM3780110 guthrie blood spot from ART conceived individual
## GSM3780111 guthrie blood spot from ART conceived individual
## data_processing
## GSM3780106 BeadStudio software v3.2
## GSM3780107 BeadStudio software v3.2
## GSM3780108 BeadStudio software v3.2
## GSM3780109 BeadStudio software v3.2
## GSM3780110 BeadStudio software v3.2
## GSM3780111 BeadStudio software v3.2
## data_processing.1
## GSM3780106 matrix processed includes: Normalized (SWAN) Average Beta and detP
## GSM3780107 matrix processed includes: Normalized (SWAN) Average Beta and detP
## GSM3780108 matrix processed includes: Normalized (SWAN) Average Beta and detP
## GSM3780109 matrix processed includes: Normalized (SWAN) Average Beta and detP
## GSM3780110 matrix processed includes: Normalized (SWAN) Average Beta and detP
## GSM3780111 matrix processed includes: Normalized (SWAN) Average Beta and detP
## data_processing.2
## GSM3780106 matrix signal intensities includes: The Methylated and unmethylated signal intensities and detP
## GSM3780107 matrix signal intensities includes: The Methylated and unmethylated signal intensities and detP
## GSM3780108 matrix signal intensities includes: The Methylated and unmethylated signal intensities and detP
## GSM3780109 matrix signal intensities includes: The Methylated and unmethylated signal intensities and detP
## GSM3780110 matrix signal intensities includes: The Methylated and unmethylated signal intensities and detP
## GSM3780111 matrix signal intensities includes: The Methylated and unmethylated signal intensities and detP
## platform_id contact_name contact_email
## GSM3780106 GPL21145 Boris,,Novakovic boris.novakovic@mcri.edu.au
## GSM3780107 GPL21145 Boris,,Novakovic boris.novakovic@mcri.edu.au
## GSM3780108 GPL21145 Boris,,Novakovic boris.novakovic@mcri.edu.au
## GSM3780109 GPL21145 Boris,,Novakovic boris.novakovic@mcri.edu.au
## GSM3780110 GPL21145 Boris,,Novakovic boris.novakovic@mcri.edu.au
## GSM3780111 GPL21145 Boris,,Novakovic boris.novakovic@mcri.edu.au
## contact_phone contact_institute
## GSM3780106 0434980073 Murdoch Children's Research Institute
## GSM3780107 0434980073 Murdoch Children's Research Institute
## GSM3780108 0434980073 Murdoch Children's Research Institute
## GSM3780109 0434980073 Murdoch Children's Research Institute
## GSM3780110 0434980073 Murdoch Children's Research Institute
## GSM3780111 0434980073 Murdoch Children's Research Institute
## contact_address
## GSM3780106 Murdoch Children's Research Institute, Royal Children's Hospital, Flemington Road
## GSM3780107 Murdoch Children's Research Institute, Royal Children's Hospital, Flemington Road
## GSM3780108 Murdoch Children's Research Institute, Royal Children's Hospital, Flemington Road
## GSM3780109 Murdoch Children's Research Institute, Royal Children's Hospital, Flemington Road
## GSM3780110 Murdoch Children's Research Institute, Royal Children's Hospital, Flemington Road
## GSM3780111 Murdoch Children's Research Institute, Royal Children's Hospital, Flemington Road
## contact_city contact_state contact_zip/postal_code contact_country
## GSM3780106 Parkville Victoria 3052 Australia
## GSM3780107 Parkville Victoria 3052 Australia
## GSM3780108 Parkville Victoria 3052 Australia
## GSM3780109 Parkville Victoria 3052 Australia
## GSM3780110 Parkville Victoria 3052 Australia
## GSM3780111 Parkville Victoria 3052 Australia
## supplementary_file
## GSM3780106 ftp://ftp.ncbi.nlm.nih.gov/geo/samples/GSM3780nnn/GSM3780106/suppl/GSM3780106_202818860092_R07C01_Grn.idat.gz
## GSM3780107 ftp://ftp.ncbi.nlm.nih.gov/geo/samples/GSM3780nnn/GSM3780107/suppl/GSM3780107_202822930094_R07C01_Grn.idat.gz
## GSM3780108 ftp://ftp.ncbi.nlm.nih.gov/geo/samples/GSM3780nnn/GSM3780108/suppl/GSM3780108_202818860162_R01C01_Grn.idat.gz
## GSM3780109 ftp://ftp.ncbi.nlm.nih.gov/geo/samples/GSM3780nnn/GSM3780109/suppl/GSM3780109_202818860162_R03C01_Grn.idat.gz
## GSM3780110 ftp://ftp.ncbi.nlm.nih.gov/geo/samples/GSM3780nnn/GSM3780110/suppl/GSM3780110_202822930046_R01C01_Grn.idat.gz
## GSM3780111 ftp://ftp.ncbi.nlm.nih.gov/geo/samples/GSM3780nnn/GSM3780111/suppl/GSM3780111_202818860100_R01C01_Grn.idat.gz
## supplementary_file.1
## GSM3780106 ftp://ftp.ncbi.nlm.nih.gov/geo/samples/GSM3780nnn/GSM3780106/suppl/GSM3780106_202818860092_R07C01_Red.idat.gz
## GSM3780107 ftp://ftp.ncbi.nlm.nih.gov/geo/samples/GSM3780nnn/GSM3780107/suppl/GSM3780107_202822930094_R07C01_Red.idat.gz
## GSM3780108 ftp://ftp.ncbi.nlm.nih.gov/geo/samples/GSM3780nnn/GSM3780108/suppl/GSM3780108_202818860162_R01C01_Red.idat.gz
## GSM3780109 ftp://ftp.ncbi.nlm.nih.gov/geo/samples/GSM3780nnn/GSM3780109/suppl/GSM3780109_202818860162_R03C01_Red.idat.gz
## GSM3780110 ftp://ftp.ncbi.nlm.nih.gov/geo/samples/GSM3780nnn/GSM3780110/suppl/GSM3780110_202822930046_R01C01_Red.idat.gz
## GSM3780111 ftp://ftp.ncbi.nlm.nih.gov/geo/samples/GSM3780nnn/GSM3780111/suppl/GSM3780111_202818860100_R01C01_Red.idat.gz
## data_row_count art status:ch1 art subtype:ch1 gender:ch1
## GSM3780106 0 ART Frozen embryo F
## GSM3780107 0 ART NA M
## GSM3780108 0 ART Fresh embryo F
## GSM3780109 0 ART Frozen embryo M
## GSM3780110 0 ART Fresh embryo F
## GSM3780111 0 ART Fresh embryo F
## sample type:ch1 time of collection:ch1
## GSM3780106 Guthrie blood spot birth
## GSM3780107 Guthrie blood spot birth
## GSM3780108 Guthrie blood spot birth
## GSM3780109 Guthrie blood spot birth
## GSM3780110 Guthrie blood spot birth
## GSM3780111 Guthrie blood spot birth
## Basename
## GSM3780106 IDAT/GSM3780106_202818860092_R07C01
## GSM3780107 IDAT/GSM3780107_202822930094_R07C01
## GSM3780108 IDAT/GSM3780108_202818860162_R01C01
## GSM3780109 IDAT/GSM3780109_202818860162_R03C01
## GSM3780110 IDAT/GSM3780110_202822930046_R01C01
## GSM3780111 IDAT/GSM3780111_202818860100_R01C01rgSet <- read.metharray.exp(targets = targets)
rgSet## class: RGChannelSet
## dim: 1051815 207
## metadata(0):
## assays(2): Green Red
## rownames(1051815): 1600101 1600111 ... 99810990 99810992
## rowData names(0):
## colnames(207): GSM3780106_202818860092_R07C01
## GSM3780107_202822930094_R07C01 ... GSM3780311_202822930170_R07C01
## GSM3780312_202822930058_R05C01
## colData names(45): title geo_accession ... Basename filenames
## Annotation
## array: IlluminaHumanMethylationEPIC
## annotation: ilm10b4.hg19mSet <- preprocessRaw(rgSet)## Loading required package: IlluminaHumanMethylationEPICmanifestmSetSw <- SWAN(mSet,verbose=FALSE)## [SWAN] Preparing normalization subset
## EPIC
## [SWAN] Normalizing methylated channel
## [SWAN] Normalizing unmethylated channelpar(mfrow=c(1,2), cex=0.8)
densityByProbeType(mSet[,1], main = "Raw")
densityByProbeType(mSetSw[,1], main = "SWAN")
Figure 1. Normalisation of bead-array data with SWAN.
# include sex chromosomes
detP <- detectionP(rgSet)
keep <- rowSums(detP < 0.01) == ncol(rgSet)
mSetSw <- mSetSw[keep,]
# exclude SNP probes
mSetSw <- mapToGenome(mSetSw)
mSetSw_nosnp <- dropLociWithSnps(mSetSw)
# exclude sex chromosomes
xyprobes <- anno$Name[anno$chr %in% c("chrX","chrY")]
mSetFlt <- mSetSw[which(!rownames(mSetSw) %in% xyprobes),]# include sex chromosomes
meth <- getMeth(mSetSw)
unmeth <- getUnmeth(mSetSw)
Mval <- log2((meth + 100)/(unmeth + 100))
beta <- getBeta(mSetSw)
dim(Mval)## [1] 810518 207# exclude sex chromosomes
meth <- getMeth(mSetFlt)
unmeth <- getUnmeth(mSetFlt)
Mval_flt <- log2((meth + 100)/(unmeth + 100))
beta_flt <- getBeta(mSetFlt)
dim(Mval_flt)## [1] 793844 207MDS analysis
par(mfrow=c(2,1))
myscree(Mval,main="incl sex chr")
myscree(Mval_flt,main="excl sex chr")
Figure 2. Screeplot shows contribution of top principal components to the overall variation in the experiment.
#MDS plot 1
colnames(targets) <- gsub(" ","_",colnames(targets))
colnames(targets) <- gsub(":","_",colnames(targets))
targets$sex<-factor(targets$`gender_ch1`)
targets$art<-factor(targets$`art_subtype`)
sample_groups <- factor(targets$art)
colour_palette=brewer.pal(n = length(levels(sample_groups)), name = "Paired")
colours <- colour_palette[as.integer(factor(targets$art))]
plot(1,axes = FALSE,xlab="",ylab="",main="MDS by ART type")
legend("center",legend=levels(sample_groups),pch=16,cex=1.2,col=colour_palette)
Figure 3. MDS plot coloured by ART classification.
mydist <- plotMDS(Mval, labels=targets$Sample_Name,col=colours,main="sex chromosomes included")
Figure 3. MDS plot coloured by ART classification.
mydist_flt <- plotMDS(Mval_flt, labels=targets$Sample_Name,col=colours,main="sex chromosomes excluded")
Figure 3. MDS plot coloured by ART classification.
colnames(mydist_flt@.Data[[5]]) <- sapply(strsplit(colnames(mydist_flt@.Data[[5]]),"_"),"[[",1)
plotMDS(mydist_flt, labels=targets$Sample_Name,col=colours,main="sex chromosomes excluded")
Figure 3. MDS plot coloured by ART classification.
#save data object
save.image("Novakovic.Rdata")#Differential analysis
non-ART Vs Fresh embryo
non-ART Vs Frozen embryo
non-ART Vs GIFT
non-ART Vs FX
GIFT Vs FX
non-ART Vs ART
Fresh Vs Frozen
non-ART Vs Fresh embryo
# sex chromosomes excluded
beta <- beta_flt
Mval <- Mval_flt
colnames(beta) <- sapply(strsplit(colnames(beta),"_"),"[[",1)
colnames(Mval) <- sapply(strsplit(colnames(Mval),"_"),"[[",1)
#targets$Basename <- gsub("IDAT/","",targets$Basename)
birth <- targets[which(targets$`time_of_collection`=="birth"),]
birth$Basename <- sapply(strsplit(birth$Basename,"/"),"[[",2)
birth$Basename <- sapply(strsplit(birth$Basename,"_"),"[[",1)
samplesheet <- subset(birth,art_subtype_ch1=="non-ART"|art_subtype_ch1=="Fresh embryo")
groups <- factor(samplesheet$art_subtype,levels=c("non-ART","Fresh embryo"))
sex <- factor(samplesheet$sex,levels=c("M","F"))
Mvals<-Mval[,colnames(Mval)%in% samplesheet$Basename]
betas<-beta[,colnames(beta)%in% samplesheet$Basename]
dim(Mvals)## [1] 793844 133dim(betas)## [1] 793844 133top_nat_vs_fh <- dm_analysis(samplesheet=samplesheet,
sex=sex,groups=groups,mx=Mvals,name="top_nat_vs_fh",
myann=myann ,beta= betas)## Your contrast returned 6294 individually significant probes. We recommend the default setting of pcutoff in dmrcate().## Fitting chr1...## Fitting chr2...## Fitting chr3...## Fitting chr4...## Fitting chr5...## Fitting chr6...## Fitting chr7...## Fitting chr8...## Fitting chr9...## Fitting chr10...## Fitting chr11...## Fitting chr12...## Fitting chr13...## Fitting chr14...## Fitting chr15...## Fitting chr16...## Fitting chr17...## Fitting chr18...## Fitting chr19...## Fitting chr20...## Fitting chr21...## Fitting chr22...## Demarcating regions...## Done!## snapshotDate(): 2023-10-24## see ?DMRcatedata and browseVignettes('DMRcatedata') for documentation## loading from cache
##
## RCircos.Core.Components initialized.
## Type ?RCircos.Reset.Plot.Parameters to see how to modify the core components.
head(top_nat_vs_fh$dma,10)## Row.names UCSC_RefGene_Name Regulatory_Feature_Group
## 743573 cg25867694 SELM Promoter_Associated
## 510348 cg16794867 AADACL2;MIR548H2
## 11452 cg00352360 RPL23;SNORA21
## 564284 cg18789663 PLD5
## 213076 cg06826756 NUP153;NUP153;NUP153
## 533323 cg17602885 CLSTN1;CLSTN1
## 656385 cg22481606 SUPT4H1 Promoter_Associated
## 457675 cg14985987 TAF1A;TAF1A Promoter_Associated
## 666703 cg22883995 CXXC5
## 181735 cg05796321 GFI1;GFI1;GFI1 Promoter_Associated
## Islands_Name Relation_to_Island logFC AveExpr
## 743573 chr22:31503236-31503871 Island -0.3197431 -3.1543540
## 510348 OpenSea -0.3196282 -0.9045549
## 11452 chr17:37009471-37010118 N_Shore -0.3853935 -2.6646947
## 564284 chr1:242687922-242688682 Island -0.3350121 -2.7477671
## 213076 chr6:17706292-17707339 S_Shore -0.2428865 -2.0644057
## 533323 OpenSea 0.3434594 5.0620765
## 656385 chr17:56429455-56429906 Island -0.3534708 -2.7252466
## 457675 chr1:222763021-222763270 S_Shore -0.2845535 -3.0255110
## 666703 chr5:139060555-139060934 N_Shore 0.5259073 3.9860142
## 181735 chr1:92945907-92952609 Island -0.2064830 -2.0441751
## t P.Value adj.P.Val B
## 743573 -6.740065 4.002684e-10 0.0002610448 12.259955
## 510348 -6.643721 6.576729e-10 0.0002610448 11.814579
## 11452 -6.362497 2.747400e-09 0.0006251414 10.532202
## 564284 -6.335282 3.149946e-09 0.0006251414 10.409565
## 213076 -6.218219 5.652691e-09 0.0006334815 9.885118
## 533323 6.199697 6.197453e-09 0.0006334815 9.802605
## 656385 -6.193526 6.390169e-09 0.0006334815 9.775143
## 457675 -6.165106 7.356488e-09 0.0006334815 9.648855
## 666703 6.133475 8.601398e-09 0.0006334815 9.508656
## 181735 -6.121734 9.114297e-09 0.0006334815 9.456718head(top_nat_vs_fh$dmr,10)## GRanges object with 10 ranges and 8 metadata columns:
## seqnames ranges strand | no.cpgs min_smoothed_fdr
## <Rle> <IRanges> <Rle> | <integer> <numeric>
## [1] chr17 4803684-4805392 * | 6 1.84800e-11
## [2] chr19 20348926-20349303 * | 5 9.66793e-13
## [3] chr13 27295928-27296010 * | 3 1.74480e-09
## [4] chr17 19265610-19266474 * | 12 1.34809e-13
## [5] chr21 40759534-40759694 * | 5 2.81859e-10
## [6] chr19 21949968-21950478 * | 8 2.88976e-11
## [7] chr15 94218934-94218949 * | 2 1.41398e-07
## [8] chr11 130298267-130298435 * | 2 3.08719e-09
## [9] chr1 247453113-247453569 * | 2 1.29713e-06
## [10] chr4 69215305-69215675 * | 3 6.31834e-13
## Stouffer HMFDR Fisher maxdiff meandiff
## <numeric> <numeric> <numeric> <numeric> <numeric>
## [1] 1.61754e-08 0.00747914 2.99196e-07 -0.1122837 -0.0676837
## [2] 2.88431e-05 0.00603051 4.29423e-05 -0.0442672 -0.0281093
## [3] 2.22231e-05 0.00897050 8.81314e-05 0.1001111 0.0966798
## [4] 3.64236e-04 0.00631189 1.58548e-04 -0.0201262 -0.0091826
## [5] 3.12521e-05 0.02859498 2.31212e-04 -0.0684461 -0.0496675
## [6] 2.17547e-03 0.01485327 3.19117e-04 -0.0242880 -0.0137829
## [7] 1.77731e-04 0.00513695 3.69585e-04 0.0315884 0.0288846
## [8] 3.15478e-04 0.00236486 4.81171e-04 -0.0393315 -0.0241258
## [9] 2.77984e-04 0.00473441 5.28762e-04 0.0276621 0.0206343
## [10] 1.39674e-03 0.00279958 5.30118e-04 -0.0191229 -0.0089473
## overlapping.genes
## <character>
## [1] C17orf107, CHRNE
## [2] CTC-260E6.6
## [3] <NA>
## [4] B9D1
## [5] WRB
## [6] ZNF100
## [7] RP11-739G5.1
## [8] ADAMTS8
## [9] <NA>
## [10] YTHDC1
## -------
## seqinfo: 22 sequences from an unspecified genome; no seqlengthstop_nat_vs_fh$comp## $up_comp
## all up OR fisherPval
## Intergenic 591632 1283 6.75888464 3.746602e-91
## Gene_Associated 1918 2 0.61310586 7.778665e-01
## Gene_Associated_Cell_type_specific 2648 6 1.33663178 4.695166e-01
## NonGene_Associated 1475 0 0.00000000 1.904277e-01
## NonGene_Associated_Cell_type_specific 263 0 0.00000000 1.000000e+00
## Promoter_Associated 102232 10 0.05046133 7.989352e-65
## Promoter_Associated_Cell_type_specific 6790 1 0.08591943 2.716676e-04
## Unclassified 39680 17 0.24242890 1.824472e-13
## Unclassified_Cell_type_specific 47206 29 0.34734851 3.218343e-11
## lowerCI upperCI
## Intergenic 5.266097571 8.8127232
## Gene_Associated 0.074118596 2.2220757
## Gene_Associated_Cell_type_specific 0.489230577 2.9208605
## NonGene_Associated 0.000000000 1.4714551
## NonGene_Associated_Cell_type_specific 0.000000000 8.3133191
## Promoter_Associated 0.024121841 0.0931117
## Promoter_Associated_Cell_type_specific 0.002177156 0.4796762
## Unclassified 0.140760498 0.3896904
## Unclassified_Cell_type_specific 0.231678604 0.5012873
##
## $dn_comp
## all dn OR fisherPval
## Intergenic 591632 1242 0.1410163 0.000000e+00
## Gene_Associated 1918 4 0.3910319 5.625372e-02
## Gene_Associated_Cell_type_specific 2648 2 0.1412284 1.444660e-04
## NonGene_Associated 1475 30 3.9110102 1.041814e-09
## NonGene_Associated_Cell_type_specific 263 1 0.7151332 1.000000e+00
## Promoter_Associated 102232 2007 6.2552095 0.000000e+00
## Promoter_Associated_Cell_type_specific 6790 77 2.1707935 2.205593e-09
## Unclassified 39680 497 2.5607872 3.621858e-68
## Unclassified_Cell_type_specific 47206 354 1.4539240 1.693842e-10
## lowerCI upperCI
## Intergenic 0.13186280 0.1507512
## Gene_Associated 0.10637301 1.0035626
## Gene_Associated_Cell_type_specific 0.01711805 0.5109711
## NonGene_Associated 2.62557566 5.6158614
## NonGene_Associated_Cell_type_specific 0.01803336 4.0212130
## Promoter_Associated 5.88248887 6.6512517
## Promoter_Associated_Cell_type_specific 1.70734718 2.7231782
## Unclassified 2.32601825 2.8145530
## Unclassified_Cell_type_specific 1.29977943 1.6222318top_nat_vs_fh$cgi## $up_comp
## all up OR fisherPval lowerCI upperCI
## Island 154463 99 0.3276677 7.638660e-36 0.2642836 0.4023450
## N_Shelf 29471 62 1.2509607 9.653741e-02 0.9531798 1.6149753
## N_Shore 76654 83 0.6134690 4.637538e-06 0.4851746 0.7665335
## OpenSea 440553 978 2.1221415 5.317916e-38 1.8807358 2.3986562
## S_Shelf 27295 57 1.2404533 1.156398e-01 0.9339052 1.6181613
## S_Shore 65408 69 0.6004154 1.202826e-05 0.4641388 0.7653312
##
## $dn_comp
## all dn OR fisherPval lowerCI upperCI
## Island 154463 3118 12.00034604 0.000000e+00 11.19530983 12.8670822
## N_Shelf 29471 28 0.17268890 2.360751e-37 0.11459367 0.2500242
## N_Shore 76654 312 0.74710572 2.898517e-07 0.66343628 0.8388190
## OpenSea 440553 456 0.09637317 0.000000e+00 0.08724113 0.1062243
## S_Shelf 27295 22 0.14666911 9.435042e-38 0.09181892 0.2224410
## S_Shore 65408 278 0.78566375 7.369766e-05 0.69300395 0.8877893names(top_nat_vs_fh)## [1] "dma" "dm_up" "dm_dn" "confects" "dmr" "comp" "cgi"
## [8] "fit"head(top_nat_vs_fh$confects$table$name)## [1] "cg06438056" "cg00397324" "cg22883995" "cg25867694" "cg00352360"
## [6] "cg16794867"saveRDS(top_nat_vs_fh,file="novakovic_nat_vs_fh.rds")non-ART Vs Frozen embryo
samplesheet<-subset(birth,art_subtype_ch1=="non-ART"|art_subtype_ch1=="Frozen embryo")
groups <- factor(samplesheet$art_subtype,levels=c("non-ART","Frozen embryo"))
sex <- factor(samplesheet$sex,levels=c("M","F"))
Mvals<-Mval[,colnames(Mval)%in% samplesheet$Basename]
betas<-beta[,colnames(beta)%in% samplesheet$Basename]
top_nat_vs_fz <- dm_analysis(samplesheet=samplesheet,
sex=sex,groups=groups,mx=Mvals,name="top_nat_vs_fz",
myann=myann ,beta= betas)
head(top_nat_vs_fz$dma,10)## Row.names UCSC_RefGene_Name Regulatory_Feature_Group
## 256518 cg08242354
## 752334 cg26210521 SLC2A9;SLC2A9
## 338484 cg10992198 WDR62;WDR62 Gene_Associated
## 462440 cg15147113 SERPINE2;SERPINE2;SERPINE2;SERPINE2 Unclassified
## 510348 cg16794867 AADACL2;MIR548H2
## 216261 cg06924355 PAQR3 Promoter_Associated
## 682894 cg23516310
## 588078 cg19738653 EGR3;EGR3;EGR3
## 675978 cg23244910
## 745548 cg25950293 ADGRE5;ADGRE5;ADGRE5
## Islands_Name Relation_to_Island logFC AveExpr
## 256518 OpenSea -0.3161459 0.8696292
## 752334 OpenSea -0.3038727 1.6980534
## 338484 OpenSea -0.2264006 1.2194271
## 462440 OpenSea -1.7913062 -2.7893219
## 510348 OpenSea -0.2958527 -0.8222307
## 216261 chr4:79860213-79860745 S_Shore -0.4309118 -3.4907576
## 682894 OpenSea -0.3822108 -3.3719970
## 588078 chr8:22547486-22553427 Island -0.4320845 -3.8315723
## 675978 chr6:106433984-106434459 Island -0.5371988 -4.0595850
## 745548 OpenSea -0.3626856 -1.1326790
## t P.Value adj.P.Val B
## 256518 -6.483935 4.327787e-09 0.003435588 9.294257
## 752334 -5.936936 5.036375e-08 0.018357258 7.280397
## 338484 -5.864166 6.937355e-08 0.018357258 7.016969
## 462440 -5.699797 1.421404e-07 0.028209332 6.426454
## 510348 -5.539701 2.834531e-07 0.044322829 5.857725
## 216261 -5.500640 3.349990e-07 0.044322829 5.719991
## 682894 -5.390687 5.346241e-07 0.060629735 5.334548
## 588078 -5.197058 1.204638e-06 0.093387142 4.664373
## 675978 -5.144676 1.497029e-06 0.093387142 4.485061
## 745548 -5.112001 1.713409e-06 0.093387142 4.373656head(top_nat_vs_fz$dmr,10)## NULLtop_nat_vs_fz$comp## NULLtop_nat_vs_fz$cgi## NULLnames(top_nat_vs_fz)## [1] "dma" "dm_up" "dm_dn" "confects" "dmr" "comp" "cgi"
## [8] "fit"head(top_nat_vs_fz$confects$table$name)## [1] "cg08242354" "cg15147113" "cg26210521" "cg10992198" "cg16794867"
## [6] "cg06924355"saveRDS(top_nat_vs_fz,file="novakovic_nat_vs_fz.rds")non-ART Vs GIFT
samplesheet<-subset(birth,art_subtype_ch1=="non-ART"|art_subtype_ch1=="GIFT")
groups <- factor(samplesheet$art_subtype,levels=c("non-ART","GIFT"))
sex <- factor(samplesheet$sex,levels=c("M","F"))
Mvals<-Mval[,colnames(Mval)%in% samplesheet$Basename]
betas<-beta[,colnames(beta)%in% samplesheet$Basename]
top_nat_vs_GIFT <- dm_analysis(samplesheet=samplesheet,
sex=sex,groups=groups,mx=Mvals,name="top_nat_vs_GIFT",
myann=myann ,beta= betas)
head(top_nat_vs_GIFT$dma,10)## Row.names UCSC_RefGene_Name
## 733585 cg25472804
## 272999 cg08792630 FOXO3;FOXO3
## 633406 cg21538355
## 534372 cg17643276 LRRC8A;LRRC8A;LRRC8A;CCBL1;CCBL1;CCBL1;CCBL1;CCBL1
## 398954 cg13101990
## 24396 cg00744351 CHRNE;C17orf107
## 452798 cg14817867 PRPSAP2
## 415481 cg13644640 NSMCE2
## 421788 cg13844500
## 325866 cg10553748 CHRNE;C17orf107
## Regulatory_Feature_Group Islands_Name
## 733585 Unclassified
## 272999 Unclassified_Cell_type_specific chr6:108878830-108883404
## 633406 chr12:38557328-38557693
## 534372 Promoter_Associated chr9:131643911-131644749
## 398954
## 24396 Promoter_Associated_Cell_type_specific chr17:4802265-4805402
## 452798
## 415481
## 421788
## 325866 Promoter_Associated_Cell_type_specific chr17:4802265-4805402
## Relation_to_Island logFC AveExpr t P.Value
## 733585 OpenSea 0.4350502 1.74435344 5.273934 8.054398e-07
## 272999 S_Shore 0.4893859 -0.24216326 5.220489 1.009636e-06
## 633406 N_Shore 0.5021382 1.28563186 5.088177 1.757637e-06
## 534372 Island -0.3625955 -2.51755908 -5.070258 1.893619e-06
## 398954 OpenSea 0.3606685 2.29037968 5.047023 2.085297e-06
## 24396 Island -1.0582572 -2.53686338 -5.036304 2.179989e-06
## 452798 OpenSea 0.3815062 1.33967837 5.017227 2.358974e-06
## 415481 OpenSea 0.3653703 2.57869751 4.992319 2.614354e-06
## 421788 OpenSea 0.5303590 1.74323790 4.970712 2.857551e-06
## 325866 Island -0.4327298 -0.07124654 -4.943129 3.200219e-06
## adj.P.Val B
## 733585 0.1869483 5.038924
## 272999 0.1869483 4.851131
## 633406 0.1869483 4.390362
## 534372 0.1869483 4.328426
## 398954 0.1869483 4.248288
## 24396 0.1869483 4.211379
## 452798 0.1869483 4.145800
## 415481 0.1869483 4.060373
## 421788 0.1869483 3.986452
## 325866 0.1869483 3.892336head(top_nat_vs_GIFT$dmr,10)## NULLtop_nat_vs_GIFT$comp## NULLtop_nat_vs_GIFT$cgi## NULLnames(top_nat_vs_GIFT)## [1] "dma" "dm_up" "dm_dn" "confects" "dmr" "comp" "cgi"
## [8] "fit"head(top_nat_vs_GIFT$confects$table$name)## [1] "cg25472804" "cg08792630" "cg21538355" "cg17643276" "cg13101990"
## [6] "cg00744351"saveRDS(top_nat_vs_GIFT,file="novakovic_nat_vs_GIFT.rds")non-ART Vs FX
samplesheet <- subset(birth,art_subtype_ch1=="non-ART"|art_subtype_ch1=="Frozen embryo" | art_subtype_ch1=="Fresh embryo")
samplesheet$art_subtype <- gsub("Fresh embryo","FX",samplesheet$art_subtype)
samplesheet$art_subtype <- gsub("Frozen embryo","FX",samplesheet$art_subtype)
groups <- factor(samplesheet$art_subtype,levels=c("non-ART","FX"))
sex <- factor(samplesheet$sex,levels=c("M","F"))
Mvals<-Mval[,colnames(Mval)%in% samplesheet$Basename]
betas<-beta[,colnames(beta)%in% samplesheet$Basename]
top_nat_vs_FX <- dm_analysis(samplesheet=samplesheet,
sex=sex,groups=groups,mx=Mvals,name="top_nat_vs_FX",
myann=myann ,beta= betas)## Your contrast returned 10346 individually significant probes. We recommend the default setting of pcutoff in dmrcate().## Fitting chr1...## Fitting chr2...## Fitting chr3...## Fitting chr4...## Fitting chr5...## Fitting chr6...## Fitting chr7...## Fitting chr8...## Fitting chr9...## Fitting chr10...## Fitting chr11...## Fitting chr12...## Fitting chr13...## Fitting chr14...## Fitting chr15...## Fitting chr16...## Fitting chr17...## Fitting chr18...## Fitting chr19...## Fitting chr20...## Fitting chr21...## Fitting chr22...## Demarcating regions...## Done!## snapshotDate(): 2023-10-24## see ?DMRcatedata and browseVignettes('DMRcatedata') for documentation## loading from cache
##
## RCircos.Core.Components initialized.
## Type ?RCircos.Reset.Plot.Parameters to see how to modify the core components.
head(top_nat_vs_FX$dma,10)## Row.names UCSC_RefGene_Name Regulatory_Feature_Group
## 510348 cg16794867 AADACL2;MIR548H2
## 656385 cg22481606 SUPT4H1 Promoter_Associated
## 564284 cg18789663 PLD5
## 11452 cg00352360 RPL23;SNORA21
## 12931 cg00397324 MX1;MX1 Promoter_Associated_Cell_type_specific
## 256518 cg08242354
## 529461 cg17465697 TRIM11 Promoter_Associated
## 363209 cg11847929 TXNDC15 Promoter_Associated
## 774705 cg27039610
## 200896 cg06438056 AK2;AK2;AK2;AK2 Promoter_Associated
## Islands_Name Relation_to_Island logFC AveExpr
## 510348 OpenSea -0.3127516 -0.9228362
## 656385 chr17:56429455-56429906 Island -0.3511758 -2.7465378
## 564284 chr1:242687922-242688682 Island -0.3250268 -2.7643316
## 11452 chr17:37009471-37010118 N_Shore -0.3688346 -2.6813081
## 12931 chr21:42798146-42798884 Island -0.5930455 -4.3802351
## 256518 OpenSea -0.2751722 0.8001948
## 529461 chr1:228593811-228594713 Island -0.4864113 -2.3689614
## 363209 chr5:134209887-134210504 Island -0.5003220 -4.0391538
## 774705 chr17:19015446-19015696 S_Shore -0.3716653 -3.3777683
## 200896 chr1:33502201-33502719 Island -0.5259693 -1.8769775
## t P.Value adj.P.Val B
## 510348 -7.111071 3.179814e-11 2.524276e-05 14.64372
## 656385 -6.799644 1.764717e-10 4.828473e-05 13.09021
## 564284 -6.793495 1.824719e-10 4.828473e-05 13.05991
## 11452 -6.643066 4.113697e-10 5.759879e-05 12.32310
## 12931 -6.642568 4.124723e-10 5.759879e-05 12.32067
## 256518 -6.632517 4.353409e-10 5.759879e-05 12.27177
## 529461 -6.375310 1.705514e-09 1.474420e-04 11.03441
## 363209 -6.369337 1.759833e-09 1.474420e-04 11.00601
## 774705 -6.340491 2.047029e-09 1.474420e-04 10.86906
## 200896 -6.324205 2.229020e-09 1.474420e-04 10.79190head(top_nat_vs_FX$dmr,10)## GRanges object with 10 ranges and 8 metadata columns:
## seqnames ranges strand | no.cpgs min_smoothed_fdr
## <Rle> <IRanges> <Rle> | <integer> <numeric>
## [1] chr17 4804104-4805392 * | 5 1.29868e-13
## [2] chr13 27295928-27296010 * | 3 3.74138e-11
## [3] chr19 21949968-21950478 * | 8 4.19106e-14
## [4] chr20 32255052-32256071 * | 4 2.18395e-10
## [5] chr1 92949337-92950836 * | 29 1.13137e-23
## [6] chr17 19265610-19266474 * | 12 8.06435e-15
## [7] chr1 3567986-3569046 * | 14 1.74179e-18
## [8] chr22 38141814-38142561 * | 10 3.53347e-15
## [9] chr12 105724039-105724818 * | 8 2.50159e-14
## [10] chr4 330865-332198 * | 14 1.28050e-13
## Stouffer HMFDR Fisher maxdiff meandiff
## <numeric> <numeric> <numeric> <numeric> <numeric>
## [1] 1.24089e-09 0.00235505 1.90034e-08 -0.1076782 -0.07216368
## [2] 1.34864e-06 0.00335990 6.27518e-06 0.1027451 0.10076823
## [3] 1.20019e-04 0.00544642 7.17078e-06 -0.0252374 -0.01380415
## [4] 1.45010e-06 0.00541807 9.11019e-06 -0.1363439 -0.08297463
## [5] 9.84886e-04 0.00196951 9.39863e-06 -0.0278868 -0.00750793
## [6] 2.70432e-05 0.00232780 9.73911e-06 -0.0187401 -0.00879585
## [7] 4.06282e-05 0.01221926 3.09056e-05 -0.0372556 -0.01337938
## [8] 3.46585e-05 0.02300114 3.75578e-05 -0.0197323 -0.01242203
## [9] 6.44131e-04 0.00666587 4.73630e-05 -0.0218061 -0.00884753
## [10] 5.22467e-04 0.02076765 5.41507e-05 -0.0120939 -0.00652470
## overlapping.genes
## <character>
## [1] C17orf107, CHRNE
## [2] <NA>
## [3] ZNF100
## [4] ACTL10, NECAB3
## [5] GFI1
## [6] B9D1
## [7] WRAP73
## [8] NOL12, TRIOBP
## [9] C12orf75
## [10] ZNF141, RP11-478C6.2
## -------
## seqinfo: 22 sequences from an unspecified genome; no seqlengthstop_nat_vs_FX$comp## $up_comp
## all up OR fisherPval
## Intergenic 591632 1494 7.01014862 6.483607e-108
## Gene_Associated 1918 3 0.79166582 1.000000e+00
## Gene_Associated_Cell_type_specific 2648 7 1.34162468 3.775519e-01
## NonGene_Associated 1475 1 0.34259488 3.819337e-01
## NonGene_Associated_Cell_type_specific 263 0 0.00000000 1.000000e+00
## Promoter_Associated 102232 11 0.04771045 5.274736e-76
## Promoter_Associated_Cell_type_specific 6790 2 0.14788379 2.841594e-04
## Unclassified 39680 16 0.19574192 6.666925e-18
## Unclassified_Cell_type_specific 47206 33 0.33979137 3.126738e-13
## lowerCI upperCI
## Intergenic 5.540407502 8.99249352
## Gene_Associated 0.162921285 2.32133562
## Gene_Associated_Cell_type_specific 0.537967476 2.77473045
## NonGene_Associated 0.008665303 1.91455601
## NonGene_Associated_Cell_type_specific 0.000000000 7.14606568
## Promoter_Associated 0.023773828 0.08563613
## Promoter_Associated_Cell_type_specific 0.017908592 0.53531665
## Unclassified 0.111585458 0.31893717
## Unclassified_Cell_type_specific 0.233005510 0.47933452
##
## $dn_comp
## all dn OR fisherPval
## Intergenic 591632 2228 0.1360485 0.000000e+00
## Gene_Associated 1918 3 0.1597442 1.932075e-05
## Gene_Associated_Cell_type_specific 2648 4 0.1541434 2.542205e-07
## NonGene_Associated 1475 47 3.3774632 4.348528e-12
## NonGene_Associated_Cell_type_specific 263 2 0.7829094 1.000000e+00
## Promoter_Associated 102232 3786 6.7671790 0.000000e+00
## Promoter_Associated_Cell_type_specific 6790 142 2.2045733 1.766074e-16
## Unclassified 39680 866 2.4420217 6.993718e-107
## Unclassified_Cell_type_specific 47206 616 1.3814880 2.316855e-13
## lowerCI upperCI
## Intergenic 0.12942460 0.1429935
## Gene_Associated 0.03289524 0.4677129
## Gene_Associated_Cell_type_specific 0.04195680 0.3952868
## NonGene_Associated 2.46788896 4.5201000
## NonGene_Associated_Cell_type_specific 0.09429758 2.8575761
## Promoter_Associated 6.46758979 7.0820030
## Promoter_Associated_Cell_type_specific 1.85081076 2.6080090
## Unclassified 2.27111284 2.6233491
## Unclassified_Cell_type_specific 1.26959406 1.5011773top_nat_vs_FX$cgi## $up_comp
## all up OR fisherPval lowerCI upperCI
## Island 154463 115 0.3273399 2.180619e-41 0.2682930 0.3960387
## N_Shelf 29471 76 1.3228757 2.262797e-02 1.0360833 1.6668838
## N_Shore 76654 100 0.6373102 4.379003e-06 0.5150645 0.7808923
## OpenSea 440553 1118 1.9992864 5.383135e-38 1.7902334 2.2359047
## S_Shelf 27295 76 1.4326854 3.457044e-03 1.1220748 1.8053330
## S_Shore 65408 82 0.6145033 5.016846e-06 0.4857653 0.7678775
##
## $dn_comp
## all dn OR fisherPval lowerCI upperCI
## Island 154463 5698 12.23270018 0.000000e+00 11.61774581 12.88535230
## N_Shelf 29471 44 0.14788841 6.517733e-73 0.10735691 0.19882000
## N_Shore 76654 618 0.81563499 7.478939e-07 0.74981741 0.88598317
## OpenSea 440553 772 0.08784057 0.000000e+00 0.08143225 0.09464214
## S_Shelf 27295 33 0.11993495 2.750645e-74 0.08246800 0.16859164
## S_Shore 65408 529 0.82077778 8.170225e-06 0.74976114 0.89707044names(top_nat_vs_FX)## [1] "dma" "dm_up" "dm_dn" "confects" "dmr" "comp" "cgi"
## [8] "fit"head(top_nat_vs_FX$confects$table$name)## [1] "cg00397324" "cg06438056" "cg11847929" "cg17465697" "cg16794867"
## [6] "cg22481606"saveRDS(top_nat_vs_FX,file="novakovic_nat_vs_FX.rds")GIFT Vs FX
samplesheet <- subset(birth,art_subtype_ch1=="GIFT"|art_subtype_ch1=="Frozen embryo" | art_subtype_ch1=="Fresh embryo")
samplesheet$art_subtype <- gsub("Fresh embryo","FX",samplesheet$art_subtype)
samplesheet$art_subtype <- gsub("Frozen embryo","FX",samplesheet$art_subtype)
groups <- factor(samplesheet$art_subtype,levels=c("GIFT","FX"))
sex <- factor(samplesheet$sex,levels=c("M","F"))
Mvals<-Mval[,colnames(Mval)%in% samplesheet$Basename]
betas<-beta[,colnames(beta)%in% samplesheet$Basename]
top_GIFT_vs_FX <- dm_analysis(samplesheet=samplesheet,
sex=sex,groups=groups,mx=Mvals,name="top_GIFT_vs_FX",
myann=myann ,beta= betas)
head(top_GIFT_vs_FX$dma,10)## Row.names UCSC_RefGene_Name
## 649276 cg22179564
## 599622 cg20185668 RBM6;RBM6
## 268646 cg08646202 ALG9;ALG9;ALG9;ALG9
## 253124 cg08129561 SLC4A8;SLC4A8;SLC4A8;SLC4A8;SLC4A8
## 13056 cg00400743 PRMT10
## 338870 cg11005961 LTN1
## 280404 cg09035199 RARB;RARB;RARB;RARB;RARB;RARB;RARB;RARB;RARB;RARB
## 430093 cg14099387 FAXDC2
## 179576 cg05723277
## 714964 cg24749153
## Regulatory_Feature_Group Islands_Name
## 649276 Unclassified_Cell_type_specific
## 599622
## 268646
## 253124
## 13056 chr4:148604954-148605625
## 338870 Promoter_Associated chr21:30364965-30365342
## 280404 NonGene_Associated
## 430093
## 179576
## 714964
## Relation_to_Island logFC AveExpr t P.Value
## 649276 OpenSea -0.3690470 -0.6244919 -5.234065 5.728277e-07
## 599622 OpenSea -0.3894483 2.7842572 -5.037692 1.384867e-06
## 268646 OpenSea -0.2906418 0.3040756 -4.899925 2.539788e-06
## 253124 OpenSea -0.5298041 2.5395224 -4.883763 2.725130e-06
## 13056 N_Shore -0.4187195 1.7047237 -4.857708 3.051873e-06
## 338870 S_Shore -0.3235771 0.3786485 -4.853476 3.108409e-06
## 280404 OpenSea -0.5626289 1.4639252 -4.822397 3.555830e-06
## 430093 OpenSea -0.5064149 2.1037502 -4.782881 4.215473e-06
## 179576 OpenSea -0.4735753 1.8157184 -4.778994 4.286428e-06
## 714964 OpenSea -0.4062668 2.2942657 -4.775335 4.354262e-06
## adj.P.Val B
## 649276 0.1785771 5.347717
## 599622 0.1785771 4.607223
## 268646 0.1785771 4.098904
## 253124 0.1785771 4.039895
## 13056 0.1785771 3.945039
## 338870 0.1785771 3.929665
## 280404 0.1785771 3.817043
## 430093 0.1785771 3.674561
## 179576 0.1785771 3.660588
## 714964 0.1785771 3.647444head(top_GIFT_vs_FX$dmr,10)## NULLtop_GIFT_vs_FX$comp## NULLtop_GIFT_vs_FX$cgi## NULLnames(top_GIFT_vs_FX)## [1] "dma" "dm_up" "dm_dn" "confects" "dmr" "comp" "cgi"
## [8] "fit"head(top_GIFT_vs_FX$confects$table$name)## [1] "cg22179564" "cg20185668" "cg08646202" "cg08129561" "cg00400743"
## [6] "cg11005961"saveRDS(top_GIFT_vs_FX,file="novakovic_GIFT_vs_FX.rds")non-ART Vs ART
samplesheet<-birth
groups <- factor(samplesheet$art_status,levels=c("non-ART","ART"))
sex <- factor(samplesheet$sex,levels=c("M","F"))
Mvals<-Mval[,colnames(Mval)%in% samplesheet$Basename]
betas<-beta[,colnames(beta)%in% samplesheet$Basename]
top_nat_vs_ART <- dm_analysis(samplesheet=samplesheet,
sex=sex,groups=groups,mx=Mvals,name="top_nat_vs_ART",
myann=myann ,beta= betas)## Your contrast returned 13365 individually significant probes. We recommend the default setting of pcutoff in dmrcate().## Fitting chr1...## Fitting chr2...## Fitting chr3...## Fitting chr4...## Fitting chr5...## Fitting chr6...## Fitting chr7...## Fitting chr8...## Fitting chr9...## Fitting chr10...## Fitting chr11...## Fitting chr12...## Fitting chr13...## Fitting chr14...## Fitting chr15...## Fitting chr16...## Fitting chr17...## Fitting chr18...## Fitting chr19...## Fitting chr20...## Fitting chr21...## Fitting chr22...## Demarcating regions...## Done!## snapshotDate(): 2023-10-24## see ?DMRcatedata and browseVignettes('DMRcatedata') for documentation## loading from cache
##
## RCircos.Core.Components initialized.
## Type ?RCircos.Reset.Plot.Parameters to see how to modify the core components.
head(top_nat_vs_ART$dma,10)## Row.names UCSC_RefGene_Name
## 656385 cg22481606 SUPT4H1
## 153960 cg04891094 GGA1;GGA1;GGA1
## 11452 cg00352360 RPL23;SNORA21
## 52374 cg01627351
## 413802 cg13592399 NID2;NID2
## 232993 cg07465122 DERL1;DERL1;DERL1;DERL1
## 195777 cg06262436 ISM1
## 45699 cg01414687 FABP3
## 226991 cg07267166 ZNF323;ZNF323;ZKSCAN3
## 564284 cg18789663 PLD5
## Regulatory_Feature_Group Islands_Name
## 656385 Promoter_Associated chr17:56429455-56429906
## 153960 Promoter_Associated chr22:38004443-38005271
## 11452 chr17:37009471-37010118
## 52374 Unclassified_Cell_type_specific
## 413802 Unclassified_Cell_type_specific chr14:52534581-52536722
## 232993 Promoter_Associated chr8:124054057-124054733
## 195777 chr20:13200670-13202616
## 45699 Unclassified
## 226991 Promoter_Associated_Cell_type_specific
## 564284 chr1:242687922-242688682
## Relation_to_Island logFC AveExpr t P.Value
## 656385 Island -0.3319894 -2.758386 -6.844132 8.171203e-11
## 153960 N_Shore -0.2840867 -3.493317 -6.746758 1.422473e-10
## 11452 N_Shore -0.3479312 -2.692333 -6.709964 1.751749e-10
## 52374 OpenSea -0.3264773 -2.626330 -6.624186 2.838683e-10
## 413802 Island -0.4458168 -3.662029 -6.617692 2.943883e-10
## 232993 Island -0.4434512 -4.562474 -6.609134 3.088398e-10
## 195777 Island -0.3611420 -2.218753 -6.570290 3.836969e-10
## 45699 OpenSea -0.4045099 -1.275311 -6.526350 4.900006e-10
## 226991 OpenSea -0.4558370 -2.137226 -6.494360 5.851119e-10
## 564284 Island -0.2878052 -2.761634 -6.478046 6.403791e-10
## adj.P.Val B
## 656385 4.086176e-05 13.86636
## 153960 4.086176e-05 13.35942
## 11452 4.086176e-05 13.16904
## 52374 4.086176e-05 12.72773
## 413802 4.086176e-05 12.69446
## 232993 4.086176e-05 12.65066
## 195777 4.351365e-05 12.45229
## 45699 4.862301e-05 12.22878
## 226991 5.083611e-05 12.06667
## 564284 5.083611e-05 11.98420head(top_nat_vs_ART$dmr,10)## GRanges object with 10 ranges and 8 metadata columns:
## seqnames ranges strand | no.cpgs min_smoothed_fdr
## <Rle> <IRanges> <Rle> | <integer> <numeric>
## [1] chr17 4803684-4805392 * | 6 4.41684e-17
## [2] chr1 92949337-92950836 * | 29 3.14221e-23
## [3] chr6 44238228-44238905 * | 4 9.69595e-13
## [4] chr19 21949968-21950478 * | 8 5.27602e-14
## [5] chr22 38141814-38142561 * | 10 2.93365e-16
## [6] chr1 150551593-150552817 * | 14 8.30981e-22
## [7] chr17 19265610-19266474 * | 12 1.60615e-14
## [8] chr13 27295928-27296010 * | 3 1.56345e-10
## [9] chr22 38597691-38599166 * | 14 8.50755e-12
## [10] chr19 53465830-53466414 * | 13 8.99240e-16
## Stouffer HMFDR Fisher maxdiff meandiff
## <numeric> <numeric> <numeric> <numeric> <numeric>
## [1] 6.95704e-13 0.000535267 1.59767e-11 -0.1055577 -0.06644571
## [2] 9.77223e-05 0.002175546 6.36421e-07 -0.0279710 -0.00746054
## [3] 4.31563e-07 0.001341528 1.40656e-06 -0.0233935 -0.02048422
## [4] 5.78744e-05 0.003658307 2.46374e-06 -0.0252468 -0.01324109
## [5] 4.98345e-06 0.012997377 4.03307e-06 -0.0197340 -0.01205356
## [6] 2.49674e-03 0.000465298 4.38951e-06 -0.0279023 -0.01012718
## [7] 7.75904e-05 0.001340427 5.21446e-06 -0.0184761 -0.00818292
## [8] 1.14592e-06 0.002987608 5.30259e-06 0.0949274 0.09400869
## [9] 2.65949e-06 0.009515391 5.32579e-06 -0.0266708 -0.01176393
## [10] 1.00915e-04 0.011836027 9.40705e-06 -0.0244057 -0.01189477
## overlapping.genes
## <character>
## [1] C17orf107, CHRNE
## [2] GFI1
## [3] TMEM151B
## [4] ZNF100
## [5] NOL12, TRIOBP
## [6] MCL1
## [7] B9D1
## [8] <NA>
## [9] MAFF, PLA2G6
## [10] ZNF816, ZNF321P, CTD..
## -------
## seqinfo: 22 sequences from an unspecified genome; no seqlengthstop_nat_vs_ART$comp## $up_comp
## all up OR fisherPval
## Intergenic 591632 3485 7.25584865 7.360445e-254
## Gene_Associated 1918 5 0.56524768 2.369424e-01
## Gene_Associated_Cell_type_specific 2648 9 0.73767229 4.697449e-01
## NonGene_Associated 1475 2 0.29356211 7.806822e-02
## NonGene_Associated_Cell_type_specific 263 0 0.00000000 6.384292e-01
## Promoter_Associated 102232 21 0.03892374 7.069511e-182
## Promoter_Associated_Cell_type_specific 6790 4 0.12665621 2.275610e-09
## Unclassified 39680 37 0.19388665 1.673507e-40
## Unclassified_Cell_type_specific 47206 87 0.38507059 1.101697e-24
## lowerCI upperCI
## Intergenic 6.20428012 8.53566549
## Gene_Associated 0.18321734 1.32261200
## Gene_Associated_Cell_type_specific 0.33663310 1.40400862
## NonGene_Associated 0.03550560 1.06305437
## NonGene_Associated_Cell_type_specific 0.00000000 3.05822384
## Promoter_Associated 0.02408602 0.05963095
## Promoter_Associated_Cell_type_specific 0.03445924 0.32471511
## Unclassified 0.13624076 0.26781369
## Unclassified_Cell_type_specific 0.30758391 0.47643477
##
## $dn_comp
## all dn OR fisherPval
## Intergenic 591632 2569 0.11585803 0.000000e+00
## Gene_Associated 1918 3 0.12371331 1.587370e-07
## Gene_Associated_Cell_type_specific 2648 2 0.05962793 4.114333e-12
## NonGene_Associated 1475 59 3.31113122 2.365917e-14
## NonGene_Associated_Cell_type_specific 263 2 0.60633982 7.774627e-01
## Promoter_Associated 102232 5031 7.29037384 0.000000e+00
## Promoter_Associated_Cell_type_specific 6790 184 2.22695715 3.310466e-21
## Unclassified 39680 1185 2.63107142 8.352424e-166
## Unclassified_Cell_type_specific 47206 871 1.53460731 1.195188e-29
## lowerCI upperCI
## Intergenic 0.110686455 0.1211931
## Gene_Associated 0.025475069 0.3621661
## Gene_Associated_Cell_type_specific 0.007220003 0.2156941
## NonGene_Associated 2.505329259 4.3010164
## NonGene_Associated_Cell_type_specific 0.073032628 2.2129215
## Promoter_Associated 7.005042284 7.5914657
## Promoter_Associated_Cell_type_specific 1.910430676 2.5822900
## Unclassified 2.472303676 2.7987174
## Unclassified_Cell_type_specific 1.428979455 1.6464108top_nat_vs_ART$cgi## $up_comp
## all up OR fisherPval lowerCI upperCI
## Island 154463 240 0.2902298 6.096625e-109 0.2535362 0.3309501
## N_Shelf 29471 183 1.3713026 6.381891e-05 1.1748638 1.5922892
## N_Shore 76654 243 0.6662687 1.158723e-10 0.5824138 0.7592553
## OpenSea 440553 2608 2.0130921 9.464542e-88 1.8726878 2.1653658
## S_Shelf 27295 172 1.3912977 5.905329e-05 1.1862373 1.6226993
## S_Shore 65408 204 0.6582494 8.439687e-10 0.5685765 0.7584508
##
## $dn_comp
## all dn OR fisherPval lowerCI upperCI
## Island 154463 7615 14.41579046 0.000000e+00 13.75949264 15.12837255
## N_Shelf 29471 46 0.11960074 6.890584e-103 0.08751775 0.15971136
## N_Shore 76654 731 0.74300193 1.220531e-15 0.68785968 0.80152112
## OpenSea 440553 802 0.06896142 0.000000e+00 0.06403268 0.07410376
## S_Shelf 27295 51 0.14371665 5.637836e-88 0.10692326 0.18919831
## S_Shore 65408 661 0.79415975 4.878765e-09 0.73251381 0.85987424names(top_nat_vs_ART)## [1] "dma" "dm_up" "dm_dn" "confects" "dmr" "comp" "cgi"
## [8] "fit"head(top_nat_vs_ART$confects$table$name)## [1] "cg00397324" "cg13592399" "cg07465122" "cg07267166" "cg11847929"
## [6] "cg17465697"saveRDS(top_nat_vs_ART,file="novakovic_nat_vs_ART.rds")Fresh embryo Vs Frozen embryo
samplesheet <- subset(birth,art_subtype_ch1=="Fresh embryo"|art_subtype_ch1=="Frozen embryo")
groups <- factor(samplesheet$art_subtype,levels=c("Fresh embryo","Frozen embryo"))
sex <- factor(samplesheet$sex,levels=c("M","F"))
Mvals<-Mval[,colnames(Mval)%in% samplesheet$Basename]
betas<-beta[,colnames(beta)%in% samplesheet$Basename]
top_fh_vs_fz <- dm_analysis(samplesheet=samplesheet,
sex=sex,groups=groups,mx=Mvals,name="top_fh_vs_fz",
myann=myann ,beta= betas)
head(top_fh_vs_fz$dma,10)## Row.names UCSC_RefGene_Name
## 84778 cg02660277 PTPRN2;PTPRN2;PTPRN2
## 741713 cg25789405
## 655472 cg22442197 RPS2;SNORA10
## 93377 cg02932204 PDZD7
## 225683 cg07224221 RTP3
## 594607 cg19983118
## 450713 cg14745383 TRAPPC9;TRAPPC9
## 297858 cg09616692 GFOD2;GFOD2
## 387340 cg12687426 KCNMB3;KCNMB3;KCNMB3;KCNMB3;KCNMB3
## 199370 cg06385817 KCTD21-AS1;KCTD21-AS1
## Regulatory_Feature_Group Islands_Name
## 84778 chr7:157440024-157442721
## 741713
## 655472 chr16:2014164-2015451
## 93377 Unclassified_Cell_type_specific
## 225683
## 594607 Promoter_Associated chr8:103822614-103823263
## 450713 Gene_Associated
## 297858
## 387340 chr3:178978948-178979364
## 199370
## Relation_to_Island logFC AveExpr t P.Value
## 84778 N_Shore -0.5837758 0.6088869 -5.099470 1.439517e-06
## 741713 OpenSea -0.4056756 3.4996145 -4.998435 2.209984e-06
## 655472 N_Shore -0.3941535 1.6464795 -4.920040 3.071840e-06
## 93377 OpenSea 0.9769342 -0.8217257 4.849491 4.120906e-06
## 225683 OpenSea -0.3849991 2.7301403 -4.848165 4.143619e-06
## 594607 Island -0.3879062 -3.1599441 -4.826148 4.539162e-06
## 450713 OpenSea -0.3504850 3.5987531 -4.818790 4.679357e-06
## 297858 OpenSea -0.4322990 2.8210026 -4.753997 6.109411e-06
## 387340 N_Shore -1.0556522 2.5173178 -4.670751 8.579195e-06
## 199370 OpenSea -0.3004069 1.6016670 -4.664075 8.814614e-06
## adj.P.Val B
## 84778 0.5306685 1.9227863
## 741713 0.5306685 1.6848404
## 655472 0.5306685 1.5017541
## 93377 0.5306685 1.3381889
## 225683 0.5306685 1.3351270
## 594607 0.5306685 1.2843275
## 450713 0.5306685 1.2673752
## 297858 0.6062399 1.1186717
## 387340 0.6751879 0.9291370
## 199370 0.6751879 0.9140145head(top_fh_vs_fz$dmr,10)## NULLtop_fh_vs_fz$comp## NULLtop_fh_vs_fz$cgi## NULLnames(top_fh_vs_fz)## [1] "dma" "dm_up" "dm_dn" "confects" "dmr" "comp" "cgi"
## [8] "fit"head(top_fh_vs_fz$confects$table$name)## [1] "cg02660277" "cg25789405" "cg22442197" "cg02932204" "cg07224221"
## [6] "cg19983118"saveRDS(top_fh_vs_fz,file="novakovic_fh_vs_fz.rds")SessionInfo
sessionInfo()## R version 4.3.2 (2023-10-31)
## Platform: x86_64-pc-linux-gnu (64-bit)
## Running under: Ubuntu 22.04.3 LTS
##
## Matrix products: default
## BLAS: /usr/lib/x86_64-linux-gnu/openblas-pthread/libblas.so.3
## LAPACK: /usr/lib/x86_64-linux-gnu/openblas-pthread/libopenblasp-r0.3.20.so; LAPACK version 3.10.0
##
## locale:
## [1] LC_CTYPE=en_US.UTF-8 LC_NUMERIC=C
## [3] LC_TIME=en_US.UTF-8 LC_COLLATE=en_US.UTF-8
## [5] LC_MONETARY=en_US.UTF-8 LC_MESSAGES=en_US.UTF-8
## [7] LC_PAPER=en_US.UTF-8 LC_NAME=C
## [9] LC_ADDRESS=C LC_TELEPHONE=C
## [11] LC_MEASUREMENT=en_US.UTF-8 LC_IDENTIFICATION=C
##
## time zone: Etc/UTC
## tzcode source: system (glibc)
##
## attached base packages:
## [1] grid parallel stats4 stats graphics grDevices utils
## [8] datasets methods base
##
## other attached packages:
## [1] IlluminaHumanMethylationEPICmanifest_0.3.0
## [2] ENmix_1.38.01
## [3] doParallel_1.0.17
## [4] qqman_0.1.9
## [5] RCircos_1.2.2
## [6] beeswarm_0.4.0
## [7] forestplot_3.1.3
## [8] abind_1.4-5
## [9] checkmate_2.3.1
## [10] reshape2_1.4.4
## [11] gplots_3.1.3
## [12] eulerr_7.0.0
## [13] GEOquery_2.70.0
## [14] RColorBrewer_1.1-3
## [15] IlluminaHumanMethylation450kmanifest_0.4.0
## [16] topconfects_1.18.0
## [17] DMRcatedata_2.20.0
## [18] ExperimentHub_2.10.0
## [19] AnnotationHub_3.10.0
## [20] BiocFileCache_2.10.1
## [21] dbplyr_2.4.0
## [22] DMRcate_2.16.1
## [23] limma_3.58.1
## [24] missMethyl_1.36.0
## [25] IlluminaHumanMethylationEPICanno.ilm10b4.hg19_0.6.0
## [26] IlluminaHumanMethylation450kanno.ilmn12.hg19_0.6.1
## [27] minfi_1.48.0
## [28] bumphunter_1.44.0
## [29] locfit_1.5-9.8
## [30] iterators_1.0.14
## [31] foreach_1.5.2
## [32] Biostrings_2.70.1
## [33] XVector_0.42.0
## [34] SummarizedExperiment_1.32.0
## [35] Biobase_2.62.0
## [36] MatrixGenerics_1.14.0
## [37] matrixStats_1.2.0
## [38] GenomicRanges_1.54.1
## [39] GenomeInfoDb_1.38.2
## [40] IRanges_2.36.0
## [41] S4Vectors_0.40.2
## [42] BiocGenerics_0.48.1
## [43] R.utils_2.12.3
## [44] R.oo_1.25.0
## [45] R.methodsS3_1.8.2
## [46] plyr_1.8.9
##
## loaded via a namespace (and not attached):
## [1] ProtGenerics_1.34.0 bitops_1.0-7
## [3] httr_1.4.7 dynamicTreeCut_1.63-1
## [5] tools_4.3.2 doRNG_1.8.6
## [7] backports_1.4.1 utf8_1.2.4
## [9] R6_2.5.1 HDF5Array_1.30.0
## [11] lazyeval_0.2.2 Gviz_1.46.1
## [13] rhdf5filters_1.14.1 permute_0.9-7
## [15] withr_2.5.2 prettyunits_1.2.0
## [17] gridExtra_2.3 base64_2.0.1
## [19] preprocessCore_1.64.0 cli_3.6.2
## [21] sass_0.4.8 readr_2.1.4
## [23] genefilter_1.84.0 askpass_1.2.0
## [25] Rsamtools_2.18.0 foreign_0.8-85
## [27] siggenes_1.76.0 illuminaio_0.44.0
## [29] dichromat_2.0-0.1 scrime_1.3.5
## [31] BSgenome_1.70.1 readxl_1.4.3
## [33] impute_1.76.0 rstudioapi_0.15.0
## [35] RSQLite_2.3.4 generics_0.1.3
## [37] BiocIO_1.12.0 gtools_3.9.5
## [39] dplyr_1.1.4 Matrix_1.6-1.1
## [41] interp_1.1-5 fansi_1.0.6
## [43] lifecycle_1.0.4 yaml_2.3.8
## [45] edgeR_4.0.3 rhdf5_2.46.1
## [47] SparseArray_1.2.2 blob_1.2.4
## [49] promises_1.2.1 crayon_1.5.2
## [51] lattice_0.22-5 GenomicFeatures_1.54.1
## [53] annotate_1.80.0 KEGGREST_1.42.0
## [55] pillar_1.9.0 knitr_1.45
## [57] beanplot_1.3.1 rjson_0.2.21
## [59] codetools_0.2-19 glue_1.6.2
## [61] data.table_1.14.10 vctrs_0.6.5
## [63] png_0.1-8 cellranger_1.1.0
## [65] gtable_0.3.4 assertthat_0.2.1
## [67] cachem_1.0.8 xfun_0.41
## [69] S4Arrays_1.2.0 mime_0.12
## [71] survival_3.5-7 statmod_1.5.0
## [73] interactiveDisplayBase_1.40.0 ellipsis_0.3.2
## [75] nlme_3.1-163 bit64_4.0.5
## [77] bsseq_1.38.0 progress_1.2.3
## [79] filelock_1.0.3 bslib_0.6.1
## [81] nor1mix_1.3-2 irlba_2.3.5.1
## [83] KernSmooth_2.23-22 rpart_4.1.21
## [85] colorspace_2.1-0 DBI_1.1.3
## [87] Hmisc_5.1-1 nnet_7.3-19
## [89] tidyselect_1.2.0 bit_4.0.5
## [91] compiler_4.3.2 curl_5.2.0
## [93] htmlTable_2.4.2 xml2_1.3.6
## [95] RPMM_1.25 DelayedArray_0.28.0
## [97] rtracklayer_1.62.0 caTools_1.18.2
## [99] scales_1.3.0 quadprog_1.5-8
## [101] rappdirs_0.3.3 stringr_1.5.1
## [103] digest_0.6.33 rmarkdown_2.25
## [105] htmltools_0.5.7 pkgconfig_2.0.3
## [107] jpeg_0.1-10 base64enc_0.1-3
## [109] sparseMatrixStats_1.14.0 highr_0.10
## [111] fastmap_1.1.1 ensembldb_2.26.0
## [113] rlang_1.1.2 htmlwidgets_1.6.4
## [115] shiny_1.8.0 DelayedMatrixStats_1.24.0
## [117] jquerylib_0.1.4 jsonlite_1.8.8
## [119] BiocParallel_1.36.0 mclust_6.0.1
## [121] VariantAnnotation_1.48.1 RCurl_1.98-1.13
## [123] magrittr_2.0.3 Formula_1.2-5
## [125] GenomeInfoDbData_1.2.11 Rhdf5lib_1.24.1
## [127] munsell_0.5.0 Rcpp_1.0.11
## [129] stringi_1.8.3 zlibbioc_1.48.0
## [131] MASS_7.3-60 org.Hs.eg.db_3.18.0
## [133] deldir_2.0-2 splines_4.3.2
## [135] multtest_2.58.0 hms_1.1.3
## [137] rngtools_1.5.2 geneplotter_1.80.0
## [139] biomaRt_2.58.0 BiocVersion_3.18.0
## [141] XML_3.99-0.16 evaluate_0.23
## [143] calibrate_1.7.7 latticeExtra_0.6-30
## [145] biovizBase_1.50.0 BiocManager_1.30.22
## [147] tzdb_0.4.0 httpuv_1.6.13
## [149] tidyr_1.3.0 openssl_2.1.1
## [151] purrr_1.0.2 reshape_0.8.9
## [153] ggplot2_3.4.4 xtable_1.8-4
## [155] restfulr_0.0.15 AnnotationFilter_1.26.0
## [157] later_1.3.2 tibble_3.2.1
## [159] memoise_2.0.1 AnnotationDbi_1.64.1
## [161] GenomicAlignments_1.38.0 cluster_2.1.4