m3Csites_S100A4_ecdf_compare.R
shuheimitsutomi
2024-07-29
suppressPackageStartupMessages(library(tidyverse))
devtools::load_all('~/Google Drive/My Drive/Scripts/R_packages/myUtilities/')## ℹ Loading myUtilitiesSettings
data_dir <- '/Volumes/Mitsu_NGS_3/METTL2A/'
wd <- "~/Google Drive/My Drive/Analysis/METTL2A/"
setwd(wd)
figdir <- paste0(wd, 'Figures/DRS_m3C_RNAs/Isoforms/Mapping_m3Csites_isoforms_ecdf_compare/')
tabledir <- paste0(wd, 'Tables/DRS_m3C_RNAs/Isoforms/')
theme_set(
theme_classic(base_size = 7) +
theme(legend.position = 'bottom')
)Functions
read_signal_parameter_files <- function(
.glob,
.dir = '/Volumes/Mitsu_NGS_2/METTL2A/Tables/Nanocompore/Signal_parameters/',
...) {
fs::dir_ls(.dir, glob = .glob) |>
read_tsv(
col_names = c('ref_pos', 'ref_kmer', 'dwell_time', 'intensity'),
col_types = 'dcdd',
id = 'filename', ...
) |>
separate(filename, sep = '_',
into = c(NA, NA, NA, NA, NA, 'sample_num', 'type', 'si', NA, 'transcript_id')) |>
mutate(transcript_id = str_remove(transcript_id, '.tsv'))
}
remove_outlier_from_eachsample <- function(df, outlier_thresh = .05) {
df |>
group_by(transcript_id, ref_pos, sample_num) |>
filter(intensity > quantile(intensity, probs = outlier_thresh / 2), na.rm = TRUE) |>
filter(intensity < quantile(intensity, probs = 1 - outlier_thresh / 2), na.rm = TRUE) |>
ungroup()
}Read data
tsv_files <-
fs::dir_ls(
'/Volumes/Mitsu_NGS_2/METTL2A/Tables/Nanocompore/Signal_parameters/',
glob = '*.tsv') |>
fs::path_filter(
regexp = 'ENST00000368716.9|ENST00000354332.8|ESPRESSO:chr1:1668:8'
)
tsv_files## /Volumes/Mitsu_NGS_2/METTL2A/Tables/Nanocompore/Signal_parameters/221117_DrTaniue_7_siMETTL2A_I_N1_ENST00000354332.8.tsv
## /Volumes/Mitsu_NGS_2/METTL2A/Tables/Nanocompore/Signal_parameters/221117_DrTaniue_7_siMETTL2A_I_N1_ENST00000368716.9.tsv
## /Volumes/Mitsu_NGS_2/METTL2A/Tables/Nanocompore/Signal_parameters/221117_DrTaniue_8_siMETTL2A_I_N2_ENST00000354332.8.tsv
## /Volumes/Mitsu_NGS_2/METTL2A/Tables/Nanocompore/Signal_parameters/221117_DrTaniue_8_siMETTL2A_I_N2_ENST00000368716.9.tsv
## /Volumes/Mitsu_NGS_2/METTL2A/Tables/Nanocompore/Signal_parameters/221117_DrTaniue_9_siMETTL2A_I_N3_ENST00000354332.8.tsv
## /Volumes/Mitsu_NGS_2/METTL2A/Tables/Nanocompore/Signal_parameters/221117_DrTaniue_9_siMETTL2A_I_N3_ENST00000368716.9.tsv
## /Volumes/Mitsu_NGS_2/METTL2A/Tables/Nanocompore/Signal_parameters/221122_DrTaniue_4_siMETTL2A_G_N1_ENST00000354332.8.tsv
## /Volumes/Mitsu_NGS_2/METTL2A/Tables/Nanocompore/Signal_parameters/221122_DrTaniue_4_siMETTL2A_G_N1_ENST00000368716.9.tsv
## /Volumes/Mitsu_NGS_2/METTL2A/Tables/Nanocompore/Signal_parameters/221122_DrTaniue_5_siMETTL2A_G_N2_ENST00000354332.8.tsv
## /Volumes/Mitsu_NGS_2/METTL2A/Tables/Nanocompore/Signal_parameters/221122_DrTaniue_5_siMETTL2A_G_N2_ENST00000368716.9.tsv
## /Volumes/Mitsu_NGS_2/METTL2A/Tables/Nanocompore/Signal_parameters/221122_DrTaniue_6_siMETTL2A_G_N3_ENST00000354332.8.tsv
## /Volumes/Mitsu_NGS_2/METTL2A/Tables/Nanocompore/Signal_parameters/221122_DrTaniue_6_siMETTL2A_G_N3_ENST00000368716.9.tsv
## /Volumes/Mitsu_NGS_2/METTL2A/Tables/Nanocompore/Signal_parameters/221128_DrTaniue_1_Cont_D_N1_ENST00000354332.8.tsv
## /Volumes/Mitsu_NGS_2/METTL2A/Tables/Nanocompore/Signal_parameters/221128_DrTaniue_1_Cont_D_N1_ENST00000368716.9.tsv
## /Volumes/Mitsu_NGS_2/METTL2A/Tables/Nanocompore/Signal_parameters/221128_DrTaniue_2_Cont_D_N2_ENST00000354332.8.tsv
## /Volumes/Mitsu_NGS_2/METTL2A/Tables/Nanocompore/Signal_parameters/221128_DrTaniue_2_Cont_D_N2_ENST00000368716.9.tsv
## /Volumes/Mitsu_NGS_2/METTL2A/Tables/Nanocompore/Signal_parameters/221128_DrTaniue_3_Cont_D_N3_ENST00000354332.8.tsv
## /Volumes/Mitsu_NGS_2/METTL2A/Tables/Nanocompore/Signal_parameters/221128_DrTaniue_3_Cont_D_N3_ENST00000368716.9.tsvparameters <-
tsv_files |>
map(read_signal_parameter_files) |>
reduce(bind_rows)
parameters## # A tibble: 20,090,588 × 8
## sample_num type si transcript_id ref_pos ref_kmer dwell_time intensity
## <chr> <chr> <chr> <chr> <dbl> <chr> <dbl> <dbl>
## 1 7 siMETTL… I ENST00000354… 7 CTCTA 0.149 77.9
## 2 7 siMETTL… I ENST00000354… 8 TCTAC 0.0113 84.7
## 3 7 siMETTL… I ENST00000354… 9 CTACA 0.0150 83.5
## 4 7 siMETTL… I ENST00000354… 10 TACAA 0.00633 81.8
## 5 7 siMETTL… I ENST00000354… 11 ACAAC 0.00267 83.5
## 6 7 siMETTL… I ENST00000354… 12 CAACC 0.0120 89.2
## 7 7 siMETTL… I ENST00000354… 13 AACCC 0.0283 83.8
## 8 7 siMETTL… I ENST00000354… 14 ACCCT 0.00733 67.9
## 9 7 siMETTL… I ENST00000354… 15 CCCTC 0.00500 73.6
## 10 7 siMETTL… I ENST00000354… 16 CCTCT 0.00233 78.7
## # ℹ 20,090,578 more rowsECDF
S100A4_TCCTG <-
parameters |>
filter(ref_kmer == 'TCCTG') |>
filter(
(transcript_id == 'ENST00000368716.9' & ref_pos == 55) |
(transcript_id == 'ENST00000354332.8' & ref_pos == 100)
)
S100A4_TCCTG |>
export_tsv(outdir = tabledir, compression = 'gz')##
## Exported to: ~/Google Drive/My Drive/Analysis/METTL2A/Tables/DRS_m3C_RNAs/Isoforms/S100A4_TCCTG_2024-07-29.tsv.gz## # A tibble: 39,628 × 8
## sample_num type si transcript_id ref_pos ref_kmer dwell_time intensity
## <chr> <chr> <chr> <chr> <dbl> <chr> <dbl> <dbl>
## 1 7 siMETTL… I ENST00000354… 100 TCCTG 0.00467 78.7
## 2 7 siMETTL… I ENST00000354… 100 TCCTG 0.00267 73.7
## 3 7 siMETTL… I ENST00000354… 100 TCCTG 0.0200 72.8
## 4 7 siMETTL… I ENST00000354… 100 TCCTG 0.00300 73.1
## 5 7 siMETTL… I ENST00000354… 100 TCCTG 0.00967 74.8
## 6 7 siMETTL… I ENST00000354… 100 TCCTG 0.0110 74.1
## 7 7 siMETTL… I ENST00000354… 100 TCCTG 0.00633 76.3
## 8 7 siMETTL… I ENST00000354… 100 TCCTG 0.0110 77.1
## 9 7 siMETTL… I ENST00000354… 100 TCCTG 0.0163 71.9
## 10 7 siMETTL… I ENST00000354… 100 TCCTG 0.0100 74.8
## # ℹ 39,618 more rowsS100A4_TCCTG_intensity_ecdf <-
parameters |>
filter(ref_kmer == 'TCCTG') |>
filter(
(transcript_id == 'ENST00000368716.9' & ref_pos == 55) |
(transcript_id == 'ENST00000354332.8' & ref_pos == 100)
) |>
remove_outlier_from_eachsample() |>
ggplot(aes(x = intensity, colour = si, linetype = transcript_id, fill = ref_kmer)) +
scale_color_manual(values = c('#8C8C8C', '#37D9CC', '#A3A3F9')) +
stat_ecdf(lwd = 1.1)
S100A4_TCCTG_intensity_ecdf |>
ggsave_multiple_formats(
outdir = figdir, width = 6, height = 4, fontsize = 7
)
S100A4_ATCCT <-
parameters |>
filter(ref_kmer == 'ATCCT') |>
filter(
(transcript_id == 'ENST00000368716.9' & ref_pos == 54) |
(transcript_id == 'ENST00000354332.8' & ref_pos == 99)
)
S100A4_ATCCT |>
export_tsv(outdir = tabledir, compression = 'gz')##
## Exported to: ~/Google Drive/My Drive/Analysis/METTL2A/Tables/DRS_m3C_RNAs/Isoforms/S100A4_ATCCT_2024-07-29.tsv.gz## # A tibble: 39,634 × 8
## sample_num type si transcript_id ref_pos ref_kmer dwell_time intensity
## <chr> <chr> <chr> <chr> <dbl> <chr> <dbl> <dbl>
## 1 7 siMETTL… I ENST00000354… 99 ATCCT 0.0307 74.6
## 2 7 siMETTL… I ENST00000354… 99 ATCCT 0.00767 71.1
## 3 7 siMETTL… I ENST00000354… 99 ATCCT 0.00733 72.3
## 4 7 siMETTL… I ENST00000354… 99 ATCCT 0.0123 72.5
## 5 7 siMETTL… I ENST00000354… 99 ATCCT 0.00700 70.9
## 6 7 siMETTL… I ENST00000354… 99 ATCCT 0.0107 73.9
## 7 7 siMETTL… I ENST00000354… 99 ATCCT 0.00300 73.3
## 8 7 siMETTL… I ENST00000354… 99 ATCCT 0.00433 76.5
## 9 7 siMETTL… I ENST00000354… 99 ATCCT 0.00267 71.9
## 10 7 siMETTL… I ENST00000354… 99 ATCCT 0.00267 72.0
## # ℹ 39,624 more rowsS100A4_ATCCT_intensity_ecdf <-
parameters |>
filter(ref_kmer == 'ATCCT') |>
filter(
(transcript_id == 'ENST00000368716.9' & ref_pos == 54) |
(transcript_id == 'ENST00000354332.8' & ref_pos == 99)
) |>
remove_outlier_from_eachsample() |>
ggplot(aes(x = intensity, colour = si, linetype = transcript_id, fill = ref_kmer)) +
scale_color_manual(values = c('#8C8C8C', '#37D9CC', '#A3A3F9')) +
stat_ecdf(lwd = 1.1)
S100A4_ATCCT_intensity_ecdf |>
ggsave_multiple_formats(
outdir = figdir, width = 6, height = 4, fontsize = 7
)
Density
S100A4_TCCTG_intensity_density <-
parameters |>
filter(ref_kmer == 'TCCTG') |>
filter(
(transcript_id == 'ENST00000368716.9' & ref_pos == 55) |
(transcript_id == 'ENST00000354332.8' & ref_pos == 100)
) |>
remove_outlier_from_eachsample(outlier_thresh = .1) |>
ggplot(aes(x = intensity, fill = si, linetype = transcript_id, colour = si)) +
scale_fill_manual(values = c('#8C8C8C', '#37D9CC', '#A3A3F9')) +
scale_color_manual(values = c('#8C8C8C', '#37D9CC', '#A3A3F9')) +
geom_density(alpha = 1/2) +
facet_wrap( ~ transcript_id, ncol = 1)
S100A4_TCCTG_intensity_density |>
ggsave_multiple_formats(
outdir = figdir, width = 6, height = 4, fontsize = 7
)## Warning in grid.Call.graphics(C_polygon, x$x, x$y, index): semi-transparency is
## not supported on this device: reported only once per page
parameters |>
filter(ref_kmer == 'TCCTG') |>
filter(
#(transcript_id == 'ENST00000368716.9' & ref_pos == 55)
(transcript_id == 'ENST00000354332.8' & ref_pos == 100)
) |>
remove_outlier_from_eachsample() |>
ggplot(aes(
x = intensity, colour = si, linetype = sample_num, fill = ref_kmer)) +
scale_color_manual(values = c('#8C8C8C', '#37D9CC', '#A3A3F9')) +
stat_ecdf(lwd = 1.1) 
Sesison info
sessioninfo::session_info()## ─ Session info ───────────────────────────────────────────────────────────────
## setting value
## version R version 4.3.1 (2023-06-16)
## os macOS Sonoma 14.5
## system aarch64, darwin20
## ui X11
## language (EN)
## collate en_US.UTF-8
## ctype en_US.UTF-8
## tz Asia/Tokyo
## date 2024-07-29
## pandoc 3.1.1 @ /Applications/RStudio.app/Contents/Resources/app/quarto/bin/tools/ (via rmarkdown)
##
## ─ Packages ───────────────────────────────────────────────────────────────────
## ! package * version date (UTC) lib source
## abind 1.4-5 2016-07-21 [1] CRAN (R 4.3.0)
## backports 1.4.1 2021-12-13 [1] CRAN (R 4.3.0)
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## bit64 4.0.5 2020-08-30 [1] CRAN (R 4.3.0)
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## carData 3.0-5 2022-01-06 [1] CRAN (R 4.3.0)
## class 7.3-22 2023-05-03 [1] CRAN (R 4.3.1)
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## xfun 0.40 2023-08-09 [1] CRAN (R 4.3.0)
## xtable 1.8-4 2019-04-21 [1] CRAN (R 4.3.0)
## yaml 2.3.7 2023-01-23 [1] CRAN (R 4.3.0)
## yardstick 1.2.0 2023-04-21 [1] CRAN (R 4.3.0)
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## [1] /Library/Frameworks/R.framework/Versions/4.3-arm64/Resources/library
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## R ── Package was removed from disk.
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