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Rep <- read.table(
here(data_dir, "Homo_sapiens_triangles.txt"),
sep = "\t",
header = FALSE
)
names(Rep) <- c(
"DelStart",
"DelEnd",
"DelLength",
"MasterRepeat",
"AlternRepeats1",
"AlternRepeats2",
"AlternRepeats3",
"AlternRepeats4"
)
MasterRepeat is a repeat with arms located close to given deletion breakpoints AlternativeRepeats1-4 are repeats where: first arm == arm1 of Master; first arm == arm2 of Master; second arm == arm1 of Master; second arm == arm2 of Master; mb_del == realised deletions (exist in MitoBreak); non_del - non realised deletions == don’t exist in MitoBreak
nrow(Rep)
[1] 880
Rep <- Rep[Rep$DelStart > 5781 &
Rep$DelStart < 16569 &
Rep$DelEnd > 5781 &
Rep$DelEnd < 16569, ]
nrow(Rep)
[1] 703
Rep$AllAlternRepeats <- paste(
Rep$AlternRepeats1,
Rep$AlternRepeats2,
Rep$AlternRepeats3,
Rep$AlternRepeats4,
sep = ","
)
Rep$AllAlternRepeats <- gsub("\\,\\[\\]\\,", ",", Rep$AllAlternRepeats)
Rep$AllAlternRepeats <- gsub("^\\[", "", Rep$AllAlternRepeats)
Rep$AllAlternRepeats <- gsub("\\]$", "", Rep$AllAlternRepeats)
Rep$CenterOfRealizedRepeats <- 0
Rep$CenterOfNonRealizedRepeats <- 0
Rep$LengthOfRealizedRepeats <- 0
Rep$LengthOfNonRealizedRepeats <- 0
Rep$StartOfRealizedRepeats <- 0
Rep$StartOfNonRealizedRepeats <- 0
Rep$EndOfRealizedRepeats <- 0
Rep$EndOfNonRealizedRepeats <- 0
for (i in (1:nrow(Rep))) {
# i = 1
# format of data to get a dataset of master and all alternative repeats for each deletion (for each line of the dataset)
temp <- Rep[i, ]
AltRep <- unlist(strsplit(temp$AllAlternRepeats, "\\]\\,\\["))
AltRep <- AltRep[AltRep != ""]
AltRep <- data.frame(AltRep)
names(AltRep) <- c("WholeLine")
if (nrow(AltRep) > 0) {
AltRep$RepeatType <- "alternative"
AltRep$WholeLine <- as.character(AltRep$WholeLine)
MasterRepeat <- as.character(temp$MasterRepeat)
MasterRepeat <- gsub("^\\[", "", MasterRepeat)
MasterRepeat <- gsub("\\]$", "", MasterRepeat)
MasterRepeat <- paste(MasterRepeat, "mb_del", sep = " ")
MasterRepeat <- data.frame(MasterRepeat)
names(MasterRepeat) <- c("WholeLine")
MasterRepeat$RepeatType <- "master"
AllRep <- rbind(MasterRepeat, AltRep)
ReturnFifth <- function(x) {
unlist(strsplit(x, " "))[5]
}
AllRep$RealisedRepeat <- apply(as.matrix(AllRep$WholeLine), 1, FUN = ReturnFifth)
ReturnFirst <- function(x) {
as.numeric(unlist(strsplit(x, " "))[1])
}
AllRep$RepStart <- apply(as.matrix(AllRep$WholeLine), 1, FUN = ReturnFirst)
ReturnSecond <- function(x) {
as.numeric(unlist(strsplit(x, " "))[2])
}
AllRep$RepEnd <- apply(as.matrix(AllRep$WholeLine), 1, FUN = ReturnSecond)
AllRep <- AllRep[AllRep$RepStart > 5781 &
AllRep$RepStart < 16569 &
AllRep$RepEnd > 5781 &
AllRep$RepEnd < 16569, ]
if (nrow(AllRep) > 0) {
AllRep$Center <- (AllRep$RepEnd - AllRep$RepStart) / 2 + AllRep$RepStart
Rep$CenterOfRealizedRepeats[i] <- mean(AllRep[AllRep$RealisedRepeat == "mb_del", ]$Center)
Rep$CenterOfNonRealizedRepeats[i] <- mean(AllRep[AllRep$RealisedRepeat == "non_del", ]$Center)
AllRep$Length <- AllRep$RepEnd - AllRep$RepStart
Rep$LengthOfRealizedRepeats[i] <- mean(AllRep[AllRep$RealisedRepeat == "mb_del", ]$Length)
Rep$LengthOfNonRealizedRepeats[i] <- mean(AllRep[AllRep$RealisedRepeat == "non_del", ]$Length)
Rep$StartOfNonRealizedRepeats[i] <- mean(AllRep[AllRep$RealisedRepeat == "non_del", ]$RepStart)
Rep$StartOfRealizedRepeats[i] <- mean(AllRep[AllRep$RealisedRepeat == "mb_del", ]$RepStart)
Rep$EndOfNonRealizedRepeats[i] <- mean(AllRep[AllRep$RealisedRepeat == "non_del", ]$RepEnd)
Rep$EndOfRealizedRepeats[i] <- mean(AllRep[AllRep$RealisedRepeat == "mb_del", ]$RepEnd)
if (i == 1) {
FinalAllRep <- AllRep
}
if (i > 1) {
FinalAllRep <- rbind(FinalAllRep, AllRep)
}
}
}
}
nrow(Rep) # 703
[1] 703
Rep <- Rep[Rep$CenterOfRealizedRepeats > 0, ]
nrow(Rep) # 643
[1] 643
Rep <- Rep[Rep$CenterOfNonRealizedRepeats > 0, ] # it means there is at least one extra (third) arm to analyze
nrow(Rep) # 643
[1] 643
Rep <- Rep[!is.na(Rep$LengthOfRealizedRepeat), ] # 618
nrow(Rep)
[1] 618
## center is a bit higher in realized repeats
wilcox.test(Rep$CenterOfRealizedRepeats,
Rep$CenterOfNonRealizedRepeats,
paired = TRUE
) # significant
Wilcoxon signed rank test with continuity correction
data: Rep$CenterOfRealizedRepeats and Rep$CenterOfNonRealizedRepeats
V = 114853, p-value = 1.506e-05
alternative hypothesis: true location shift is not equal to 0
t.test(Rep$CenterOfRealizedRepeats,
Rep$CenterOfNonRealizedRepeats,
paired = TRUE
) # significant
Paired t-test
data: Rep$CenterOfRealizedRepeats and Rep$CenterOfNonRealizedRepeats
t = 4.619, df = 617, p-value = 4.696e-06
alternative hypothesis: true mean difference is not equal to 0
95 percent confidence interval:
162.6072 403.1435
sample estimates:
mean difference
282.8754
summary(Rep$CenterOfRealizedRepeats)
Min. 1st Qu. Median Mean 3rd Qu. Max.
8590 10634 11334 11500 12409 15647
summary(Rep$CenterOfNonRealizedRepeats)
Min. 1st Qu. Median Mean 3rd Qu. Max.
7021 10292 11290 11217 12196 15889
boxplot(Rep$CenterOfRealizedRepeats,
Rep$CenterOfNonRealizedRepeats,
notch = TRUE,
names = c(
"CenterOfRealizedRepeats",
"CenterOfNonRealizedRepeats"
)
)
RepReal <- Rep %>%
select(
Realized = CenterOfRealizedRepeats,
NonRealized = CenterOfNonRealizedRepeats
) %>%
gather(., Realized, NonRealized,
key = "Deletion",
value = "CenterOfRepeats"
)
pltViolRepCenterRelease <- ggstatsplot::ggbetweenstats(
data = RepReal,
x = Deletion,
y = CenterOfRepeats,
notch = TRUE,
# show notched box plot
mean.ci = TRUE,
# whether to display confidence interval for means
k = 5,
# number of decimal places for statistical results
# outlier.tagging = TRUE, # whether outliers need to be tagged
# outlier.label = ContactZone, # variable to be used for the outlier tag
xlab = "Realisation of deletion",
# label for the x-axis variable
ylab = "Center of Repeats",
# label for the y-axis variable
title = "The effect of repeats' position on deletion realisation",
# title text for the plot
ggtheme = ggthemes::theme_fivethirtyeight(),
# choosing a different theme
ggstatsplot.layer = FALSE,
# turn off `ggstatsplot` theme layer
package = "wesanderson",
# package from which color palette is to be taken
palette = "Royal1",
# choosing a different color palette
messages = TRUE
)
# Note: Shapiro-Wilk Normality Test for Center of Repeats: p-value = 0.003
# Note: Bartlett's test for homogeneity of variances for factor Realisation of deletion: p-value = < 0.001
cowplot::save_plot(
plot = pltViolRepCenterRelease,
base_height = 8,
base_asp = 1.618,
file = normalizePath(
file.path(plots_dir, "violin_rep_center_release.pdf")
)
)
pltViolRepCenterRelease
## deletion is longer in realized repeats
wilcox.test(
Rep$LengthOfRealizedRepeats,
Rep$LengthOfNonRealizedRepeats,
paired = TRUE
) # significant
Wilcoxon signed rank test with continuity correction
data: Rep$LengthOfRealizedRepeats and Rep$LengthOfNonRealizedRepeats
V = 175286, p-value < 2.2e-16
alternative hypothesis: true location shift is not equal to 0
t.test(
Rep$LengthOfRealizedRepeats,
Rep$LengthOfNonRealizedRepeats,
paired = TRUE
) # significant
Paired t-test
data: Rep$LengthOfRealizedRepeats and Rep$LengthOfNonRealizedRepeats
t = 24.481, df = 617, p-value < 2.2e-16
alternative hypothesis: true mean difference is not equal to 0
95 percent confidence interval:
1837.842 2158.412
sample estimates:
mean difference
1998.127
summary(Rep$LengthOfRealizedRepeats) # 5643 mean
Min. 1st Qu. Median Mean 3rd Qu. Max.
18 4420 5501 5643 7066 9682
summary(Rep$LengthOfNonRealizedRepeats) # 3450 Median 3645 mean
Min. 1st Qu. Median Mean 3rd Qu. Max.
12 2767 3450 3645 4489 8950
summary(Rep$LengthOfRealizedRepeats - Rep$LengthOfNonRealizedRepeats) # 1998 mean
Min. 1st Qu. Median Mean 3rd Qu. Max.
-4890 585 2116 1998 3335 7416
boxplot(Rep$LengthOfRealizedRepeats,
Rep$LengthOfNonRealizedRepeats,
notch = TRUE,
names = c(
"LengthOfRealizedRepeats",
"LengthOfNonRealizedRepeats"
)
)
RepReal <-
Rep %>%
select(
Realized = LengthOfRealizedRepeats,
NonRealized = LengthOfNonRealizedRepeats
) %>%
gather(., Realized, NonRealized,
key = "Deletion",
value = "DistanceBetweenRepeats"
) %>%
full_join(., RepReal)
pltViolRepFlankLengthRelease <-
ggstatsplot::ggbetweenstats(
data = RepReal,
x = Deletion,
y = DistanceBetweenRepeats,
notch = TRUE,
# show notched box plot
mean.ci = TRUE,
# whether to display confidence interval for means
k = 5,
# number of decimal places for statistical results
# outlier.tagging = TRUE, # whether outliers need to be tagged
# outlier.label = ContactZone, # variable to be used for the outlier tag
xlab = "Realisation of deletion",
# label for the x-axis variable
ylab = "Distance between Repeats",
# label for the y-axis variable
title = "The effect of repeats' distance on deletion realisation",
# title text for the plot
ggtheme = ggthemes::theme_fivethirtyeight(),
# choosing a different theme
ggstatsplot.layer = FALSE,
# turn off `ggstatsplot` theme layer
package = "wesanderson",
# package from which color palette is to be taken
palette = "Royal1",
# choosing a different color palette
messages = TRUE
)
# Note: Bartlett's test for homogeneity of variances for factor Realisation of deletion: p-value = < 0.001
cowplot::save_plot(
plot = pltViolRepFlankLengthRelease,
base_height = 8,
base_asp = 1.618,
file = normalizePath(
file.path(plots_dir, "violin_rep_flanked_length_release.pdf")
)
)
pltViolRepFlankLengthRelease
## start and end in realized versus non-realized repeats
wilcox.test(Rep$StartOfRealizedRepeats,
Rep$StartOfNonRealizedRepeats,
paired = TRUE
) # significant
Wilcoxon signed rank test with continuity correction
data: Rep$StartOfRealizedRepeats and Rep$StartOfNonRealizedRepeats
V = 44922, p-value < 2.2e-16
alternative hypothesis: true location shift is not equal to 0
t.test(Rep$StartOfRealizedRepeats,
Rep$StartOfNonRealizedRepeats,
paired = TRUE
) # significant
Paired t-test
data: Rep$StartOfRealizedRepeats and Rep$StartOfNonRealizedRepeats
t = -9.2587, df = 617, p-value < 2.2e-16
alternative hypothesis: true mean difference is not equal to 0
95 percent confidence interval:
-868.0955 -564.2809
sample estimates:
mean difference
-716.1882
summary(Rep$StartOfRealizedRepeats) # mean 8678: 25%: 7401 - 75% 9764:
Min. 1st Qu. Median Mean 3rd Qu. Max.
5810 7401 8472 8678 9764 15638
quantile(Rep$StartOfRealizedRepeats, 0.1) # 6465
10%
6465
quantile(Rep$StartOfRealizedRepeats, 0.9) # 10954
90%
10954
summary(Rep$StartOfNonRealizedRepeats) # mean 9394
Min. 1st Qu. Median Mean 3rd Qu. Max.
5873 8330 9112 9394 10434 15718
summary(Rep$StartOfNonRealizedRepeats - Rep$StartOfRealizedRepeats) # mean 716 => non realised start later (~700 bp): 9394 - 8678
Min. 1st Qu. Median Mean 3rd Qu. Max.
-6137.0 -303.1 393.0 716.2 1797.4 7416.5
wilcox.test(Rep$EndOfRealizedRepeats, Rep$EndOfNonRealizedRepeats, paired = TRUE) # significant
Wilcoxon signed rank test with continuity correction
data: Rep$EndOfRealizedRepeats and Rep$EndOfNonRealizedRepeats
V = 125093, p-value < 2.2e-16
alternative hypothesis: true location shift is not equal to 0
t.test(Rep$EndOfRealizedRepeats, Rep$EndOfNonRealizedRepeats, paired = TRUE) # significant
Paired t-test
data: Rep$EndOfRealizedRepeats and Rep$EndOfNonRealizedRepeats
t = 18.41, df = 617, p-value < 2.2e-16
alternative hypothesis: true mean difference is not equal to 0
95 percent confidence interval:
1145.196 1418.682
sample estimates:
mean difference
1281.939
summary(Rep$EndOfRealizedRepeats) # 14321 mean
Min. 1st Qu. Median Mean 3rd Qu. Max.
10970 13696 14124 14321 15362 16274
summary(Rep$EndOfNonRealizedRepeats) # 13039 mean
Min. 1st Qu. Median Mean 3rd Qu. Max.
7451 12132 13270 13039 13992 16261
summary(Rep$EndOfNonRealizedRepeats - Rep$EndOfRealizedRepeats) # mean -1281=> non realised end earlier (~1300 bp): 13039 - 14321
Min. 1st Qu. Median Mean 3rd Qu. Max.
-7334.5 -2198.6 -934.2 -1281.9 0.0 4890.0
quantile(Rep$EndOfRealizedRepeats, 0.1) # 13286
10%
13286.6
quantile(Rep$EndOfRealizedRepeats, 0.9) # 15863
90%
15863.4
plot(
FinalAllRep[FinalAllRep$RealisedRepeat == "non_del", ]$RepStart,
FinalAllRep[FinalAllRep$RealisedRepeat == "non_del", ]$RepEnd,
pch = 16,
col = "grey",
xlim = c(5781, 16569),
ylim = c(16569, 5781),
xlab = "",
ylab = ""
)
par(new = TRUE)
plot(
FinalAllRep[FinalAllRep$RealisedRepeat == "mb_del", ]$RepStart,
FinalAllRep[FinalAllRep$RealisedRepeat == "mb_del", ]$RepEnd,
pch = 16,
col = "red",
xlim = c(5781, 16569),
ylim = c(16569, 5781),
xlab = "Start",
ylab = "End"
)
# Visualise difference between density distributions of realized vs non-realized repeats.
sp <- ggplot(
FinalAllRep,
aes(
RepStart,
RepEnd
)
) +
aes(colour = RealisedRepeat) +
geom_point() +
scale_y_reverse() +
theme_minimal(17) +
xlab("5’") +
ylab("3’") +
scale_x_continuous(breaks = c(15000, 12000, 9000, 6000), position = "bottom") +
scale_color_manual(
values = c("red", "grey"),
labels = c(
"Realized repeats",
"Non-realized repeats"
)
) +
theme(
legend.position = "bottom",
legend.title = element_blank(),
legend.box = "horizontal"
)
gg_realised <-
ggExtra::ggMarginal(
sp,
type = "density",
margins = "both",
size = 3,
groupColour = TRUE,
groupFill = TRUE
)
cowplot::save_plot(here(plots_dir, "real-vs-nonreal-repeats.svg"), gg_realised, base_asp = 0.868, base_height = 7)
wilcox.test(FinalAllRep[FinalAllRep$RealisedRepeat == "mb_del", ]$RepStart, FinalAllRep[FinalAllRep$RealisedRepeat == "non_del", ]$RepStart)
Wilcoxon rank sum test with continuity correction
data: FinalAllRep[FinalAllRep$RealisedRepeat == "mb_del", ]$RepStart and FinalAllRep[FinalAllRep$RealisedRepeat == "non_del", ]$RepStart
W = 1354700, p-value = 1.299e-07
alternative hypothesis: true location shift is not equal to 0
wilcox.test(FinalAllRep[FinalAllRep$RealisedRepeat == "mb_del", ]$RepEnd, FinalAllRep[FinalAllRep$RealisedRepeat == "non_del", ]$RepEnd)
Wilcoxon rank sum test with continuity correction
data: FinalAllRep[FinalAllRep$RealisedRepeat == "mb_del", ]$RepEnd and FinalAllRep[FinalAllRep$RealisedRepeat == "non_del", ]$RepEnd
W = 2151564, p-value < 2.2e-16
alternative hypothesis: true location shift is not equal to 0
FinalAllRep$RealisedRepeat <- as.factor(FinalAllRep$RealisedRepeat)
summary(FinalAllRep$RealisedRepeat)
mb_del non_del
902 3391
StartA <- 6000 # 7000
StartB <- 9000 # 10000
EndA <- 13000
EndB <- 16000
R.In <- nrow(Rep[Rep$StartOfRealizedRepeats >= StartA &
Rep$StartOfRealizedRepeats <= StartB &
Rep$EndOfRealizedRepeats >= EndA &
Rep$EndOfRealizedRepeats <= EndB, ])
R.Out <- nrow(Rep[(Rep$StartOfRealizedRepeats < StartA |
Rep$StartOfRealizedRepeats > StartB) |
(Rep$EndOfRealizedRepeats < EndA |
Rep$EndOfRealizedRepeats > EndB), ])
R.In + R.Out # 324 + 294 = 618
[1] 618
NR.In <- nrow(Rep[Rep$StartOfNonRealizedRepeats >= StartA &
Rep$StartOfNonRealizedRepeats <= StartB &
Rep$EndOfNonRealizedRepeats >= EndA &
Rep$EndOfNonRealizedRepeats <= EndB, ])
NR.Out <- nrow(Rep[(Rep$StartOfNonRealizedRepeats < StartA |
Rep$StartOfNonRealizedRepeats > StartB) |
(Rep$EndOfNonRealizedRepeats < EndA |
Rep$EndOfNonRealizedRepeats > EndB), ])
NR.In + NR.Out # 79 + 598 = 618
[1] 618
X <- cbind(c(R.In, R.Out), c(NR.In, NR.Out))
fisher.test(X) # odds 7.5, p-value < 2.2e-16
Fisher's Exact Test for Count Data
data: X
p-value < 2.2e-16
alternative hypothesis: true odds ratio is not equal to 1
95 percent confidence interval:
5.615152 10.113074
sample estimates:
odds ratio
7.505642
Visualize it as mosaic plot
X <- data.frame(X)
names(X) <- c("R", "NR")
row.names(X) <- c("IN", "OUT")
mosaicplot(X, main = "")
gg_realised
Visualise difference between density distributions of realized vs non-realized repeats.
sessionInfo()
R version 4.2.2 (2022-10-31)
Platform: x86_64-pc-linux-gnu (64-bit)
Running under: Ubuntu 22.04.1 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
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
attached base packages:
[1] stats graphics grDevices datasets utils methods base
other attached packages:
[1] ggpubr_0.6.0 here_1.0.1 lubridate_1.9.2 forcats_1.0.0
[5] stringr_1.5.0 dplyr_1.1.1 purrr_1.0.1 readr_2.1.4
[9] tidyr_1.3.0 tibble_3.2.1 ggplot2_3.4.2 tidyverse_2.0.0
[13] workflowr_1.7.0
loaded via a namespace (and not attached):
[1] fs_1.6.1 insight_0.19.1 httr_1.4.5
[4] rprojroot_2.0.3 tools_4.2.2 backports_1.4.1
[7] bslib_0.4.2 utf8_1.2.3 R6_2.5.1
[10] statsExpressions_1.5.0 colorspace_2.1-0 withr_2.5.0
[13] tidyselect_1.2.0 processx_3.8.0 compiler_4.2.2
[16] git2r_0.31.0 textshaping_0.3.6 performance_0.10.2
[19] cli_3.6.1 prismatic_1.1.1 labeling_0.4.2
[22] bayestestR_0.13.0 sass_0.4.5 scales_1.2.1
[25] mvtnorm_1.1-3 callr_3.7.3 pbapply_1.7-0
[28] systemfonts_1.0.4 digest_0.6.31 svglite_2.1.1
[31] rmarkdown_2.21 pkgconfig_2.0.3 htmltools_0.5.5
[34] fastmap_1.1.1 highr_0.10 ggthemes_4.2.4
[37] rlang_1.1.0 rstudioapi_0.14 shiny_1.7.4
[40] farver_2.1.1 jquerylib_0.1.4 generics_0.1.3
[43] jsonlite_1.8.4 car_3.1-2 magrittr_2.0.3
[46] parameters_0.20.2 patchwork_1.1.2 Matrix_1.5-3
[49] Rcpp_1.0.10 munsell_0.5.0 fansi_1.0.4
[52] abind_1.4-5 lifecycle_1.0.3 stringi_1.7.12
[55] whisker_0.4.1 yaml_2.3.7 carData_3.0-5
[58] grid_4.2.2 paletteer_1.5.0 ggrepel_0.9.3
[61] parallel_4.2.2 promises_1.2.0.1 miniUI_0.1.1.1
[64] lattice_0.20-45 cowplot_1.1.1 hms_1.1.3
[67] zeallot_0.1.0 knitr_1.42 ps_1.7.4
[70] pillar_1.9.0 ggsignif_0.6.4 effectsize_0.8.3
[73] glue_1.6.2 evaluate_0.20 getPass_0.2-2
[76] renv_0.17.2 vctrs_0.6.1 tzdb_0.3.0
[79] httpuv_1.6.9 MatrixModels_0.5-1 gtable_0.3.3
[82] BayesFactor_0.9.12-4.4 rematch2_2.1.2 datawizard_0.7.1
[85] cachem_1.0.7 ggExtra_0.10.0 xfun_0.38
[88] mime_0.12 xtable_1.8-4 correlation_0.8.3
[91] broom_1.0.4 coda_0.19-4 rstatix_0.7.2
[94] later_1.3.0 ragg_1.2.5 ggstatsplot_0.11.0
[97] timechange_0.2.0 ellipsis_0.3.2