library(optparse) library(qtl) library(stringi) library(stringr) tmp_dir = Sys.getenv("TMPDIR") option_list = list( make_option(c("-g", "--geno"), type="character", help=".geno file containing a dataset's genotypes"), make_option(c("-p", "--pheno"), type="character", help="File containing two columns - sample names and values"), make_option(c("-c", "--addcovar"), action="store_true", default=NULL, help="Use covariates (included as extra columns in the phenotype input file)"), make_option(c("--covarstruct"), type="character", help="File detailing which covariates are categorical or numerical"), make_option(c("--model"), type="character", default="normal", help="Mapping Model - Normal or Non-Parametric"), make_option(c("--method"), type="character", default="hk", help="Mapping Method - hk (Haley Knott), ehk (Extended Haley Knott), mr (Marker Regression), em (Expectation-Maximization), imp (Imputation)"), make_option(c("-i", "--interval"), action="store_true", default=NULL, help="Use interval mapping"), make_option(c("--nperm"), type="integer", default=0, help="Number of permutations"), make_option(c("--pstrata"), action="store_true", default=NULL, help="Use permutation strata (stored as final column/vector in phenotype input file)"), make_option(c("-s", "--scale"), type="character", default="mb", help="Mapping scale - Megabases (Mb) or Centimorgans (cM)"), make_option(c("--control"), type="character", default=NULL, help="Name of marker (contained in genotype file) to be used as a control"), make_option(c("-o", "--outdir"), type="character", default=file.path(tmp_dir, "output"), help="Directory in which to write result file"), make_option(c("-f", "--filename"), type="character", default=NULL, help="Name to use for result file"), make_option(c("-v", "--verbose"), action="store_true", default=NULL, help="Show extra information") ); opt_parser = OptionParser(option_list=option_list); opt = parse_args(opt_parser); verbose_print <- function(...){ if (!is.null(opt$verbose)) { for(item in list(...)){ cat(item) } cat("\n") } } adjustXprobs <- function(cross){ sex <- getsex(cross)$sex pr <- cross$geno[["X"]]$prob stopifnot(!is.null(pr), !is.null(sex)) for(i in 1:ncol(pr)) { pr[sex==0,i,3:4] <- 0 pr[sex==1,i,1:2] <- 0 pr[,i,] <- pr[,i,]/rowSums(pr[,i,]) } cross$geno[["X"]]$prob <- pr invisible(cross) } if (is.null(opt$geno) || is.null(opt$pheno)){ print_help(opt_parser) stop("Both a genotype and phenotype file must be provided.", call.=FALSE) } geno_file = opt$geno pheno_file = opt$pheno # Generate randomized filename for cross object cross_file = file.path(tmp_dir, "cross", paste(stri_rand_strings(1, 8), ".cross", sep = "")) trim <- function( x ) { gsub("(^[[:space:]]+|[[:space:]]+$)", "", x) } get_geno_code <- function(header, name = 'unk'){ mat = which(unlist(lapply(header,function(x){ length(grep(paste('@',name,sep=''), x)) })) == 1) return(trim(strsplit(header[mat],':')[[1]][2])) } geno_to_csvr <- function(genotypes, trait_names, trait_vals, out, type, sex = NULL, mapping_scale = "Mb", verbose = FALSE){ # Assume a geno header is not longer than 40 lines header = readLines(genotypes, 40) # Major hack to skip the geno headers toskip = which(unlist(lapply(header, function(x){ length(grep("Chr\t", x)) })) == 1) - 1 type <- get_geno_code(header, 'type') # Get the genotype codes if(type == '4-way'){ genocodes <- NULL } else { genocodes <- c(get_geno_code(header, 'mat'), get_geno_code(header, 'het'), get_geno_code(header, 'pat')) } genodata <- read.csv(genotypes, sep='\t', skip=toskip, header=TRUE, na.strings=get_geno_code(header,'unk'), colClasses='character', comment.char = '#') verbose_print('Genodata:', toskip, " ", dim(genodata), genocodes, '\n') # If there isn't a sex phenotype, treat all as males if(is.null(sex)) sex <- rep('m', (ncol(genodata)-4)) cross_items = list() # Add trait and covar phenotypes for (i in 1:length(trait_names)){ cross_items[[i]] <- c(trait_names[i], '', '', unlist(trait_vals[[i]])) } # Sex phenotype for the mice cross_items[[length(trait_names) + 1]] <- c('sex', '', '', sex) # Genotypes cross_items[[length(trait_names) + 2]] <- cbind(genodata[,c('Locus','Chr', mapping_scale)], genodata[, 5:ncol(genodata)]) out_csvr <- do.call(rbind, cross_items) # Save it to a file write.table(out_csvr, file=out, row.names=FALSE, col.names=FALSE, quote=FALSE, sep=',') # Load the created cross file using R/qtl read.cross if (type == '4-way') { verbose_print('Loading in as 4-WAY\n') cross = read.cross(file=out, 'csvr', genotypes=NULL, crosstype="4way") } else if(type == 'f2') { verbose_print('Loading in as F2\n') cross = read.cross(file=out, 'csvr', genotypes=genocodes, crosstype="f2") } else { verbose_print('Loading in as normal\n') cross = read.cross(file=out, 'csvr', genotypes=genocodes) } if (type == 'riset') { # If its a RIL, convert to a RIL in R/qtl verbose_print('Converting to RISELF\n') cross <- convert2riself(cross) } return(cross) } create_marker_covars <- function(the_cross, control_marker){ #' Given a string of one or more marker names (comma separated), fetch #' the markers' values from the genotypes and return them as vectors/a vector #' of values # In case spaces are added in the string of marker names covariate_names <- strsplit(str_replace(control_marker, " ", ""), ",") genotypes <- pull.geno(the_cross) covariates_in_geno <- which(covariate_names %in% colnames(genotypes)) covariate_names <- covariate_names[covariates_in_geno] marker_covars <- genotypes[, unlist(covariate_names)] return(marker_covars) } # Get phenotype vector from input file df <- read.table(pheno_file, na.strings = "x", header=TRUE, check.names=FALSE) sample_names <- df$Sample trait_names <- colnames(df)[2:length(colnames(df))] # Since there will always only be one non-covar phenotype, its name will be in the first column pheno_name = unlist(trait_names)[1] trait_vals <- vector(mode = "list", length = length(trait_names)) for (i in 1:length(trait_names)) { this_trait <- trait_names[i] this_vals <- df[this_trait] trait_vals[[i]] <- this_vals trait_names[i] = paste("T_", this_trait, sep = "") } # Get type of genotypes, since it needs to be checked before calc.genoprob header = readLines(geno_file, 40) type <- get_geno_code(header, 'type') verbose_print('Generating cross object\n') cross_object = geno_to_csvr(geno_file, trait_names, trait_vals, cross_file, type) # Calculate genotype probabilities if (!is.null(opt$interval)) { verbose_print('Calculating genotype probabilities with interval mapping\n') cross_object <- calc.genoprob(cross_object, step=5, stepwidth="max") } else { verbose_print('Calculating genotype probabilities\n') cross_object <- calc.genoprob(cross_object) } # If 4way, adjust X chromosome genotype probabilities if (type == "4-way") { verbose_print('Adjusting genotype probabilities for 4way cross') cross_object <- adjustXprobs(cross_object) } # Pull covariates out of cross object, if they exist covars <- c() # Holds the covariates which should be passed to R/qtl if (!is.null(opt$addcovar)) { # If perm strata are being used, it'll be included as the final column in the phenotype file if (!is.null(opt$pstrata)) { covar_names = trait_names[2:(length(trait_names)-1)] } else { covar_names = trait_names[2:length(trait_names)] } covars <- pull.pheno(cross_object, covar_names) # Read in the covar description file covarDescr <- read.table(opt$covarstruct, sep="\t", header=FALSE) for(x in 1:nrow(covarDescr)){ cat(covarDescr[x, 1]) name <- paste0("T_", covarDescr[x, 1]) # The covar description file doesn't have T_ in trait names (the cross object does) type <- covarDescr[x, 2] if(type == "categorical"){ if(length(table(covars[,name])) > 2){ # More then 2 levels create the model matrix for the factor mdata <- data.frame(toExpand = as.factor(covars[, name])) options(na.action='na.pass') modelmatrix <- model.matrix(~ toExpand + 0, mdata)[,-1] covars <- cbind(covars, modelmatrix) }else{ # 2 levels? just bind the trait as covar verbose_print('Binding covars to covars\n') covars <- cbind(covars, covars[,name]) } } } } # Pull permutation strata out of cross object, if it is being used perm_strata = vector() if (!is.null(opt$pstrata)) { strata_col = trait_names[length(trait_names)] perm_strata <- pull.pheno(cross_object, strata_col) } # If a marker name is supplied as covariate, get its vector of values and add them as a covariate if (!is.null(opt$control)) { marker_covars = create_marker_covars(cross_object, opt$control) covars <- cbind(covars, marker_covars) } # Calculate permutations if (opt$nperm > 0) { if (!is.null(opt$filename)){ perm_out_file = file.path(opt$outdir, paste("PERM_", opt$filename, sep = "" )) } else { perm_out_file = file.path(opt$outdir, paste(pheno_name, "_PERM_", stri_rand_strings(1, 8), sep = "")) } if (!is.null(opt$addcovar) || !is.null(opt$control)){ if (!is.null(opt$pstrata)) { verbose_print('Running ', opt$nperm, ' permutations with cofactors and strata\n') perm_results = scanone(cross_object, pheno.col=1, addcovar=covars, n.perm=opt$nperm, perm.strata=perm_strata, model=opt$model, method=opt$method) } else { verbose_print('Running ', opt$nperm, ' permutations with cofactors\n') perm_results = scanone(cross_object, pheno.col=1, addcovar=covars, n.perm=opt$nperm, model=opt$model, method=opt$method) } } else { if (!is.null(opt$pstrata)) { verbose_print('Running ', opt$nperm, ' permutations with strata\n') perm_results = scanone(cross_object, pheno.col=1, n.perm=opt$nperm, perm.strata=perm_strata, model=opt$model, method=opt$method) } else { verbose_print('Running ', opt$nperm, ' permutations\n') perm_results = scanone(cross_object, pheno.col=1, n.perm=opt$nperm, model=opt$model, method=opt$method) } } write.csv(perm_results, perm_out_file) } if (!is.null(opt$filename)){ out_file = file.path(opt$outdir, opt$filename) } else { out_file = file.path(opt$outdir, paste(pheno_name, "_", stri_rand_strings(1, 8), sep = "")) } if (!is.null(opt$addcovar) || !is.null(opt$control)){ verbose_print('Running scanone with cofactors\n') qtl_results = scanone(cross_object, pheno.col=1, addcovar=covars, model=opt$model, method=opt$method) } else { verbose_print('Running scanone\n') qtl_results = scanone(cross_object, pheno.col=1, model=opt$model, method=opt$method) } #QTL main effects on adjusted longevity getEffects <- function(sdata, gtsprob, marker = "1_24042124", model = "longevity ~ sex + site + cohort + treatment", trait = "longevity"){ rownames(sdata) <- 1:nrow(sdata) rownames(gtsprob) <- 1:nrow(gtsprob) mp <- gtsprob[, grep(marker, colnames(gtsprob))] gts <- unlist(lapply(lapply(lapply(apply(mp,1,function(x){which(x > 0.85)}),names), strsplit, ":"), function(x){ if(length(x) > 0){ return(x[[1]][2]); }else{ return(NA) } })) ismissing <- which(apply(sdata, 1, function(x){any(is.na(x))})) if(length(ismissing) > 0){ sdata <- sdata[-ismissing, ] gts <- gts[-ismissing] } mlm <- lm(as.formula(model), data = sdata) pheAdj <- rep(NA, nrow(sdata)) adj <- residuals(mlm) + mean(sdata[, trait]) pheAdj[as.numeric(names(adj))] <- adj means <- c(mean(pheAdj[which(gts == "AC")],na.rm=TRUE),mean(pheAdj[which(gts == "AD")],na.rm=TRUE),mean(pheAdj[which(gts == "BC")],na.rm=TRUE),mean(pheAdj[which(gts == "BD")],na.rm=TRUE)) std <- function(x) sd(x,na.rm=TRUE)/sqrt(length(x)) stderrs <- c(std(pheAdj[which(gts == "AC")]),std(pheAdj[which(gts == "AD")]),std(pheAdj[which(gts == "BC")]),std(pheAdj[which(gts == "BD")])) paste0(round(means,0), " ± ", round(stderrs,2)) } if (type == "4-way") { verbose_print("Get phenotype name + genoprob + all phenotypes + models for 4-way crosses") traitname <- colnames(pull.pheno(cross_object))[1] gtsp <- pull.genoprob(cross_object) allpheno <- pull.pheno(cross_object) if (!is.null(opt$addcovar)) { model <- paste0(traitname, " ~ ", paste0(covar_names, sep="", collapse=" + ")) } else { model <- paste0(traitname, " ~ 1 ") } meffects <- c() verbose_print("Getting QTL main effects for 4-way crosses") for(marker in rownames(qtl_results)){ meff <- getEffects(allpheno, gtsp, marker = marker, model, trait = traitname) meffects <- rbind(meffects, meff) } qtl_results <- cbind(data.frame(qtl_results[,1:3]), meffects) colnames(qtl_results)[4:7] <- c("AC", "AD", "BC", "BD") } write.csv(qtl_results, out_file)