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267 lines
10 KiB
267 lines
10 KiB
gs_export_pop<-function(gs, pop, trim.channels=T){
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pop.dt<-c()
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# Initializes the progress bar
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pb <- txtProgressBar(min = 0, # Minimum value of the progress bar
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max = length(gs), # Maximum value of the progress bar
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style = 3, # Progress bar style (also available style = 1 and style = 2)
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width = 50, # Progress bar width. Defaults to getOption("width")
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char = "=") # Character used to create the bar
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for (i in 1:length(gs)){
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samp<-sampleNames(gs)[i]
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data<-as.data.frame(flowCore::exprs(gh_pop_get_data(gs[[samp]], pop))) %>% add_column("Sample"=samp)
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if (length(pop.dt) > 0){
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pop.dt<-rbind(pop.dt, data)
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}else{
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pop.dt<-data
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}
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setTxtProgressBar(pb, i)
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}
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close(pb)
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if (trim.channels){
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colnames(pop.dt)<-gsub(" [A-Za-z0-9.-]*","",colnames(pop.dt))
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pop.dt<-pop.dt[colnames(pop.dt)[!colnames(pop.dt) %in% c("FSC-H","SSC-H","FSC-Width","FVS")]]
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}
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return(pop.dt)
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}
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get_cut<-function(x, markers, ref, thr){
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x<-x %>%
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dplyr::filter(Condition == ref) %>%
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gather(Marker, Value, all_of(markers)) %>%
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group_by(`ID.SAR-EM`, Marker, Tiempo) %>%
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summarise(cut=predict_cut(Value, thr))
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}
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get_positive<-function(x, cut=NULL, markers, ref, thr){
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if (is.null(cut)){
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cut<-get_cut(cd8, markers = markers, ref = ref, thr = thr)
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}
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x<-merge(
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x %>% gather(Marker, Value, all_of(unique(cut$Marker))),
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cut
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)
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x<-x %>%
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mutate(Marker_cut=Value >= cut)
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x<-x %>%
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select(`ID.SAR-EM`, Tiempo, Condition, CellID, Marker, Marker_cut) %>%
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mutate(Marker_pos=paste0(Marker,".",Marker_cut)) %>%
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group_by(`ID.SAR-EM`,Tiempo, Condition, CellID) %>%
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summarise(Pheno=paste0(Marker_pos, collapse = " ")) %>%
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# mutate(Pheno=paste0("IFNg.",IFNg_p," ","TNFa.",TNFa_p," ","CD69.",CD69_p," ","CD107a.",CD107a_p)) %>%
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mutate(Pheno=gsub("[A-Za-z0-9]*.FALSE |[A-Za-z0-9]*.FALSE$| [A-Za-z0-9]*.FALSE","",Pheno),
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Pheno=gsub(".TRUE","+",Pheno),
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Pheno=gsub("^$","Neg",Pheno))
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x.dt<-x %>%
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group_by(`ID.SAR-EM`, Tiempo, Condition, Pheno) %>%
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count() %>%
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group_by(`ID.SAR-EM`, Tiempo, Condition) %>%
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mutate(perc=n*100/sum(n))
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return(x.dt)
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}
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transform_gs<-function(gs, saveparams=NULL, trans_params_obj=NULL, index=1){
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if (is.null(trans_params_obj)){
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log_mark<-colnames(gs)[!grepl("FS|SS|Time", colnames(gs))]
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trans_params<-lapply(log_mark, function(x) list(channel=x, scale="biexp",maxvalue=250000,pos=4, widthBasis=-400, max=5, min=0))
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names(trans_params)<-log_mark
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log_mark_lin<-colnames(gs)[grepl("FS|SS|Time", colnames(gs))]
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trans_params_lin<-lapply(log_mark_lin, function(x) list(channel=x, scale="lin",
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max=max(flowCore::exprs(gh_pop_get_data(gs, "root"))[,x])*1.1, min=0))
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names(trans_params_lin)<-log_mark_lin
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trans_params<-c(trans_params_lin, trans_params)
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}else{
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log_mark<-colnames(gs)[!grepl("FSC|SSC|Time", colnames(gs))]
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log_mark_lin<-colnames(gs)[grepl("FSC|SSC|Time", colnames(gs))]
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}
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shiny::runApp(shiny::shinyApp(
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ui = fluidPage(
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sidebarLayout(
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sidebarPanel(selectInput("channel", "Channel", colnames(gs), multiple = F),
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uiOutput("maxvalue"),
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uiOutput("pos"),
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uiOutput("widthBasis"),
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uiOutput("max"),
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uiOutput("min"),
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actionButton("applyBut", "Apply"),
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actionButton("stopBut", "Quit")),
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mainPanel(plotOutput("hist"))
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)
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),
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server = function(input, output) {
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output$hist <- renderPlot({
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gs.trans<-gs_clone(gs[index])
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max<-input$max
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if (trans_params[[input$channel]]$scale == "biexp"){
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trans.obj<-flowjo_biexp_trans(maxValue = input$maxvalue, pos=input$pos,
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widthBasis=input$widthBasis)
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transList <- transformerList(input$channel, trans.obj)
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gs.trans <- transform(gs.trans, transList)
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max<-10^input$max
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}
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ggcyto(gs.trans, aes(x = .data[[input$channel]]), subset = "root")+
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geom_density(fill = "blue", alpha= 0.5)+
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ggcyto_par_set(limits = list(x = c(input$min,max)))
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})
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output$maxvalue<-renderUI({if(trans_params[[input$channel]]$scale == "biexp"){
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sliderInput("maxvalue", "maxValue", min=0, max=1000000, value=trans_params[[input$channel]]$maxvalue)}})
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output$pos<-renderUI({if(trans_params[[input$channel]]$scale == "biexp"){
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sliderInput("pos", "pos", min=0, max=10, value=trans_params[[input$channel]]$pos, step = 0.5)}})
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output$widthBasis<-renderUI({if(trans_params[[input$channel]]$scale == "biexp"){
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sliderInput("widthBasis", "widthBasis", min=-1000, max=0, value=trans_params[[input$channel]]$widthBasis)}})
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output$max<-renderUI({
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if(trans_params[[input$channel]]$scale == "biexp"){
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sliderInput("max", "Max X", min=0, max=6, value=log10(trans_params[[input$channel]]$max), step=0.1)
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}else{
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sliderInput("max", "Max X", min=0, max=trans_params[[input$channel]]$max,
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value=trans_params[[input$channel]]$max)
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}
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})
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output$min<-renderUI({
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if(trans_params[[input$channel]]$scale == "biexp"){
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sliderInput("min", "Min X", min=-1000, max=100000, value=trans_params[[input$channel]]$min)
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}else{
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sliderInput("min", "Min X", min=0, max=trans_params[[input$channel]]$max, value=trans_params[[input$channel]]$min)
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}
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})
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observeEvent(input$applyBut, {
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print("Transformation and Scaling Updated!")
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if (isolate(input$channel) %in% log_mark){
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trans_params[[isolate(input$channel)]]$maxvalue<<-isolate(input$maxvalue)
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trans_params[[isolate(input$channel)]]$pos<<-isolate(input$pos)
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trans_params[[isolate(input$channel)]]$widthBasis<<-isolate(input$widthBasis)
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trans_params[[isolate(input$channel)]]$max<<-10^isolate(input$max)
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trans_params[[isolate(input$channel)]]$min<<-isolate(input$min)
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}
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if (isolate(input$channel) %in% log_mark_lin){
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trans_params[[isolate(input$channel)]]$max<<-isolate(input$max)
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trans_params[[isolate(input$channel)]]$min<<-isolate(input$min)
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}
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})
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observeEvent(input$stopBut, {
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stopApp()
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})
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}
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))
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trans_apply(gs, trans_params)
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print("Transformation Applied")
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if (!is.null(saveparams)){
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saveRDS(trans_params, file=saveparams)
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}
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return(trans_params)
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}
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trans_apply<-function(gs, trans_params){
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for (i in 1:length(trans_params)){
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if (trans_params[[i]]$scale == "biexp"){
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print(trans_params[[i]]$channel)
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trans.obj<-flowjo_biexp_trans(maxValue = trans_params[[i]]$maxvalue,
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pos=trans_params[[i]]$pos,
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widthBasis=trans_params[[i]]$widthBasis)
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transList <- transformerList(trans_params[[i]]$channel, trans.obj)
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gs <- transform(gs, transList)
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}
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}
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}
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ggcyto_trans<-function(gs, x, y, trans_params_obj=trans_params, subset, gates=T, stats=T, ...){
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g<-ggcyto(gs, aes(.data[[x]], .data[[y]]), subset=subset, ...)+
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ggcyto_par_set(limits = list(x = c(trans_params_obj[[x]]$min,trans_params_obj[[x]]$max),
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y=c(trans_params_obj[[y]]$min,trans_params_obj[[y]]$max)))
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if(gates==T){
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if (subset == "root"){
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gates<-gs_get_pop_paths(gs)[grepl(paste0("^/","[^/]*$"), gs_get_pop_paths(gs))]
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}else{
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gates<-gs_get_pop_paths(gs)[grepl(paste0(subset,"/","[^/]*$"), gs_get_pop_paths(gs))]
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}
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g<-g+geom_point(size=0.1, alpha=0.1)+
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geom_gate(gates)
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if (stats== T){
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g<-g+geom_stats(gates)
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}
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}
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return(g)
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}
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ggcyto_trans_all<-function(gs, index=1, trans_params_obj=trans_params, stats=T,...){
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subsets<-unlist(sapply(gs_get_pop_paths(gs), function(x) strsplit(x, "/")[[1]][length(strsplit(x, "/")[[1]])-1]))
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subsets<-c("root",unique(subsets[subsets != ""]))
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g.list<-list()
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for (subset in subsets){
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if (subset == "root"){
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gates<-gs_get_pop_paths(gs)[grepl(paste0("^/","[^/]*$"), gs_get_pop_paths(gs))]
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}else{
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gates<-gs_get_pop_paths(gs)[grepl(paste0(subset,"/","[^/]*$"), gs_get_pop_paths(gs))]
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}
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subset.x<-names(gs_pop_get_gate(gs, gates[1])[[1]]@parameters)[1]
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subset.y<-names(gs_pop_get_gate(gs, gates[1])[[1]]@parameters)[2]
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g.list[[subset]]<-ggcyto::as.ggplot(ggcyto_trans(gs[[index]], subset.x, subset.y,
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trans_params_obj, subset, stats=stats))
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}
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g<-do.call(ggpubr::ggarrange, c(g.list, ...))
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g<-ggpubr::annotate_figure(g, top = ggpubr::text_grob(sampleNames(gs)[index],
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color = "black", face = "bold", size = 14))
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return(g)
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}
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LMD2FCS<-function(files, output.dir=NULL){
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if(is.null(output.dir)){
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route<-paste0(gsub("/[^/]*$","",files[1]),"/")
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}
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for (lmd in files){
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lmd<-gsub(".*/","",lmd)
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fcs<-read.FCS(paste0(route,lmd), dataset = 2)
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keyword(fcs)['$FIL']<-paste0(gsub(".LMD","",lmd), ".fcs")
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write.FCS(fcs, paste0(route, gsub(".LMD","",lmd), ".fcs"))
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}
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print("Conversión completada")
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}
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gates_save<-function(gs, file="gates.rds", save=T){
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gates.list<-list()
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pop_paths<-gs_get_pop_paths(gs)[gs_get_pop_paths(gs) != "root"]
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pop_paths<-lapply(pop_paths, function(x) rev(rev(strsplit(x, "/")[[1]])[1:2]))
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for (gate in pop_paths){
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parent<-if (gate[1] == ""){"root"}else{gate[1]}
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gated_pop<-gate[2]
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gates.list[[paste0(parent,"/",gated_pop)]]<-gs_pop_get_gate(gs, gated_pop)
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}
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if(save){saveRDS(gates.list, file)}
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return(gates.list)
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}
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gates_apply<-function(gs, gates, exact=T){
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if (exact){
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for (gate in names(gates)){
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parent<-strsplit(gate, "/")[[1]][1]
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gated_pop<-strsplit(gate, "/")[[1]][2]
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gs_pop_add(gs, gates[[gate]], parent=parent, name=gated_pop)
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print(paste0("Gate ",gated_pop," applied!"))
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}
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}else{
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for (gate in names(gates)){
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parent<-strsplit(gate, "/")[[1]][1]
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gated_pop<-strsplit(gate, "/")[[1]][2]
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gs_pop_add(gs, gates[[gate]][[1]], parent=parent, name=gated_pop)
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print(paste0("Gate ",gated_pop," applied!"))
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}
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}
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recompute(gs)
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return(gs)
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}
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