Differential methylation analysis between RA and control samples (only consider methylation on chrY in male samples)
Junyan Lu
Last updated: 2023-09-04
Checks: 5 1
Knit directory: RA_Tcell_omics/analysis/
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Load libraries
Global variables
Load and preprocess datasets
Load omics data
Only keep probes on X chromose
How many samples
Control RA
3 2 Only 3 controls and 2 RA samples. The results may be unreliable
Get unique symbol
Remove CpGs not associated with known genes
Correlation between average methylation level and phenotype
Two Sample t-test
data: meanMethylation by group
t = -1.5878, df = 3, p-value = 0.2105
alternative hypothesis: true difference in means between group Control and group RA is not equal to 0
95 percent confidence interval:
-0.06167598 0.02061802
sample estimates:
mean in group Control mean in group RA
0.4510691 0.4715981 On the overall methylation level, there’s no strong trend that RA samples have higher methylation.
PCA analysis
Calculate PCA
PCA plots
PC1 versus PC2
PC2 versus PC3
Associate PCs with disease
# A tibble: 5 × 3
# Groups: pc [5]
pc estimate p.value
<chr> <dbl> <dbl>
1 PC1 -1.95e+ 0 0.211
2 PC5 4.38e-17 0.217
3 PC2 9.23e- 1 0.372
4 PC4 4.77e- 1 0.444
5 PC3 -3.10e- 1 0.663The first three principal components can separate RA with control samples, to some degree.
Differentially methylated CpGs
Prepare data
Process methylation dataset
[1] 34 5Differential methylation using limma
Add mean difference of beta values
Save the full table as excel file
Number of hypermethylation and hypomethylation
Visualize results
associated probes
Only 1 probe
Visualize the significant associations
R version 4.2.0 (2022-04-22)
Platform: x86_64-apple-darwin17.0 (64-bit)
Running under: macOS Big Sur/Monterey 10.16
Matrix products: default
BLAS: /Library/Frameworks/R.framework/Versions/4.2/Resources/lib/libRblas.0.dylib
LAPACK: /Library/Frameworks/R.framework/Versions/4.2/Resources/lib/libRlapack.dylib
locale:
[1] en_US.UTF-8/en_US.UTF-8/en_US.UTF-8/C/en_US.UTF-8/en_US.UTF-8
attached base packages:
[1] stats4 stats graphics grDevices utils datasets methods
[8] base
other attached packages:
[1] forcats_0.5.1 stringr_1.4.1
[3] dplyr_1.0.9 purrr_0.3.4
[5] readr_2.1.2 tidyr_1.2.0
[7] tibble_3.1.8 ggplot2_3.4.1
[9] tidyverse_1.3.2 pheatmap_1.0.12
[11] SummarizedExperiment_1.26.1 Biobase_2.56.0
[13] GenomicRanges_1.48.0 GenomeInfoDb_1.32.2
[15] IRanges_2.30.0 S4Vectors_0.34.0
[17] BiocGenerics_0.42.0 MatrixGenerics_1.8.1
[19] matrixStats_0.62.0 limma_3.52.2
loaded via a namespace (and not attached):
[1] ggbeeswarm_0.6.0 googledrive_2.0.0 colorspace_2.0-3
[4] ellipsis_0.3.2 rprojroot_2.0.3 XVector_0.36.0
[7] fs_1.5.2 rstudioapi_0.13 farver_2.1.1
[10] DT_0.23 ggrepel_0.9.1 bit64_4.0.5
[13] AnnotationDbi_1.58.0 fansi_1.0.3 lubridate_1.8.0
[16] xml2_1.3.3 splines_4.2.0 cachem_1.0.6
[19] knitr_1.39 jsonlite_1.8.3 workflowr_1.7.0
[22] broom_1.0.0 annotate_1.74.0 dbplyr_2.2.1
[25] png_0.1-7 compiler_4.2.0 httr_1.4.3
[28] backports_1.4.1 assertthat_0.2.1 Matrix_1.5-4
[31] fastmap_1.1.0 gargle_1.2.0 cli_3.4.1
[34] later_1.3.0 htmltools_0.5.4 tools_4.2.0
[37] gtable_0.3.0 glue_1.6.2 GenomeInfoDbData_1.2.8
[40] Rcpp_1.0.9 cellranger_1.1.0 jquerylib_0.1.4
[43] vctrs_0.5.2 Biostrings_2.64.0 writexl_1.4.0
[46] crosstalk_1.2.0 xfun_0.31 rvest_1.0.2
[49] lifecycle_1.0.3 XML_3.99-0.10 googlesheets4_1.0.0
[52] zlibbioc_1.42.0 scales_1.2.0 hms_1.1.1
[55] promises_1.2.0.1 RColorBrewer_1.1-3 yaml_2.3.5
[58] memoise_2.0.1 sass_0.4.2 stringi_1.7.8
[61] RSQLite_2.2.15 highr_0.9 genefilter_1.78.0
[64] rlang_1.0.6 pkgconfig_2.0.3 bitops_1.0-7
[67] evaluate_0.15 lattice_0.20-45 htmlwidgets_1.5.4
[70] labeling_0.4.2 cowplot_1.1.1 bit_4.0.4
[73] tidyselect_1.1.2 magrittr_2.0.3 R6_2.5.1
[76] generics_0.1.3 DelayedArray_0.22.0 DBI_1.1.3
[79] pillar_1.8.0 haven_2.5.0 withr_2.5.0
[82] survival_3.4-0 KEGGREST_1.36.3 RCurl_1.98-1.7
[85] modelr_0.1.8 crayon_1.5.2 utf8_1.2.2
[88] tzdb_0.3.0 rmarkdown_2.14 grid_4.2.0
[91] readxl_1.4.0 blob_1.2.3 git2r_0.30.1
[94] reprex_2.0.1 digest_0.6.30 xtable_1.8-4
[97] httpuv_1.6.6 munsell_0.5.0 beeswarm_0.4.0
[100] vipor_0.4.5 bslib_0.4.1