###Assess complexity under varying parameters
###NB: The paths are relative to the root of the scphaser git repo
###
###SYNOPSIS
###library('devtools')
###devtools::load_all()
###source("ignore/code/analysis/mousehybrid/seqdepth2ngenes.R")
##Params
sys = 'rs13'
sys = 'dna'
##*###
##Dirs
##*###
if(sys == 'dna'){
cloud.dir = '/mnt/kauffman/edsgard/cloud/btsync/work/rspd'
}
if(sys == 'rs13'){
cloud.dir = '/Volumes/Data/cloud/btsync/work/rspd'
}
prefilt_rds_dir = '../nogit/data/mousehybrid/all_snps/downsampled'
##OUT
out_rds_dir = '../nogit/data/mousehybrid'
out_pdf_dir = './ignore/res/mousehybrid/perf/pdf'
##*###
##Files
##*###
##OUT
param2ngenes_rds = file.path(out_rds_dir, 'seqdepth2ngenes.rds')
##Libs
source('./ignore/code/analysis/performance.R')
library('BiocParallel')
library('dplyr')
library('tidyr')
main <- function(){
##read data
load('../nogit/data/mousehybrid/mousehybrid.rda')
##create acset
featdata = mousehybrid$featdata
ref = featdata[, 'c57']
alt = featdata[, 'cast']
featdata = cbind(featdata, ref, alt, stringsAsFactors = FALSE)
acset = new_acset(featdata, mousehybrid$refcount, mousehybrid$altcount, mousehybrid$phenodata)
lapply(acset, dim) #167,443 x 336
length(unique(acset[['featdata']][, 'feat'])) #11,590
##Filter vars with 0 counts
acset = filter_zerorow(acset)
lapply(acset, dim) #167,443
length(unique(acset[['featdata']][, 'feat'])) #11,590
##Call gt
min_acount = 3
fc = 3 #75/25
acset = call_gt(acset, min_acount, fc)
##Filter variants on number of cells where ASE towards the same allele (as to remove probable false positive calls)
alpha = 0.1
mono.ase = 0.1
if(!(mono.ase == 0)){
acset = filter_homovars(acset, alpha = alpha, mono.ase = mono.ase)
}
lapply(acset, dim) #104,933 x 336
##*###
##Randomly sample n counts
##*###
altcount = acset[['altcount']]
refcount = acset[['refcount']]
totcount = altcount + refcount
##number of reads
ntot = sum(totcount)
ntot #139,192,303
ncount = c(seq(25e6, 125e6, 25e6), ntot)
##permutation iterations
npermiter = 10 #10
perm_iter = 1:npermiter
##specify paramset as all possible combinations of the params
paramset = expand.grid(perm_iter, ncount, stringsAsFactors = FALSE)
colnames(paramset) = c('perm_iter', 'ncount')
##parallelization
ncores = 80
bp_param = BiocParallel::MulticoreParam(workers = ncores)
##Filter vars and feats
nparamset = nrow(paramset)
acset_list = BiocParallel::bplapply(1:nparamset, filter_acset_par, BPPARAM = bp_param, paramset = paramset, acset = acset)
##status: fin (11.30 -> 11.40, 80 cores)
##get number of genes
ngenes = unlist(lapply(acset_list, function(j.acset){length(unique(j.acset[['featdata']][, 'feat']))}))
param2ngenes = cbind(paramset, ngenes)
##get number of variants
nvars = unlist(lapply(acset_list, function(j.acset){nrow(j.acset[['featdata']])}))
param2ngenes = cbind(param2ngenes, nvars)
##get fraction of genes
n.bg.genes = 20268 ##see log.sh
frac.genes = param2ngenes[, 'ngenes'] / n.bg.genes
param2ngenes = cbind(param2ngenes, frac.genes)
##add col mean number of reads per cell
n.cells = ncol(altcount)
print(n.cells)
ncount.percell = param2ngenes[, 'ncount'] / n.cells
param2ngenes = cbind(param2ngenes, ncount.percell)
##Dump
saveRDS(param2ngenes, file = param2ngenes_rds)
##status: fin
summary(param2ngenes[, 'ngenes'])
##*###
##Plot
##*###
param2ngenes = readRDS(param2ngenes_rds)
library('ggplot2')
errbar.w = 2.5e6 / 336
gg = ggplot(param2ngenes, aes_string(x = 'ncount.percell', y = 'ngenes'))
gg = gg + geom_line(stat = 'summary', fun.y = 'mean')
gg = gg + geom_errorbar(stat = 'summary', fun.data = mean_se, width = errbar.w) #width = errbar.w
##tick breaks
##gg = gg + coord_cartesian(ylim = c(800, 3050))
##gg = gg + scale_y_continuous(breaks = seq(1000, 3000, 500))
##gg = gg + scale_x_continuous(breaks = c(seq(25e6, 125e6, 25e6)))
##Background
gg = gg + theme(panel.grid.major = element_blank(), panel.grid.minor = element_blank(), axis.line = element_line(colour = "black")) + theme(panel.background = element_blank())
gg = gg + theme(axis.text = element_text(colour="black", size = 15), axis.ticks = element_line(colour = 'black'), axis.title = element_text(size = 15))
gg = gg + xlab('Mean number of allele counts per cell')
gg = gg + ylab('Number of phasable genes')
gg = gg + theme(axis.line.x = element_line(color="black", size = 0.5), axis.line.y = element_line(color="black", size = 0.5))
pdf.h = 5
pdf.w = 5
j.pdf = file.path(out_pdf_dir, 'nallelecount2ngenes.pdf')
dir.create(dirname(j.pdf), recursive = TRUE)
pdf(j.pdf, height = pdf.h, width = pdf.w)
plot(gg)
dev.off()
gg = ggplot(param2ngenes, aes_string(x = 'ncount.percell', y = 'frac.genes'))
gg = gg + geom_line(stat = 'summary', fun.y = 'mean')
##tick breaks
##gg = gg + coord_cartesian(ylim = c(800, 3050))
##gg = gg + scale_y_continuous(breaks = seq(1000, 3000, 500))
##gg = gg + scale_x_continuous(breaks = c(seq(50, 336, 50), 336))
##Background
gg = gg + theme(panel.grid.major = element_blank(), panel.grid.minor = element_blank(), axis.line = element_line(colour = "black")) + theme(panel.background = element_blank())
gg = gg + theme(axis.text = element_text(colour="black", size = 15), axis.ticks = element_line(colour = 'black'), axis.title = element_text(size = 15))
gg = gg + xlab('Mean number of allele counts per cell')
gg = gg + ylab('Number of phasable genes')
gg = gg + theme(axis.line.x = element_line(color="black", size = 0.5), axis.line.y = element_line(color="black", size = 0.5))
pdf.h = 5
pdf.w = 5
j.pdf = file.path(out_pdf_dir, 'nallelecount2fracgenes.pdf')
dir.create(dirname(j.pdf), recursive = TRUE)
pdf(j.pdf, height = pdf.h, width = pdf.w)
plot(gg)
dev.off()
}
filter_acset_par <- function(j.param, paramset, acset, nmincells = 5, nminvar = 2){
j_ncount = paramset[j.param, 'ncount']
altcount = acset[['altcount']]
refcount = acset[['refcount']]
totcount = altcount + refcount
ncount = sum(totcount)
frac.down = j_ncount / ncount
##keep relative frequency btw alt and ref (skip sampling in this step)
alt.frac = sum(altcount) / ncount
ref.frac = 1 - alt.frac
alt.frac.down = alt.frac * frac.down
ref.frac.down = ref.frac * frac.down
##downsample
altcount.down = downsample.matrix(altcount, alt.frac.down)
refcount.down = downsample.matrix(refcount, ref.frac.down)
##set to downsampled data
acset[['refcount']] = refcount.down
acset[['altcount']] = altcount.down
##Call gt
min_acount = 3
fc = 3 #75/25
acset = call_gt(acset, min_acount, fc)
##filter vars and genes
acset_filt = filter_acset(acset, nmincells, nminvar)
return(acset_filt)
}
downsample.matrix <- function(count.mat, frac.down, pseudo.count = 0){
##keep relative frequency between cols (skip sampling in this step)
col2ncount = apply(count.mat, 2, sum)
col2ncount.down = ceiling(col2ncount * frac.down)
##add pseudocount (skip, since we are down-sampling there is no need to do this)
count.mat = count.mat + pseudo.count
col2ncount = apply(count.mat, 2, sum)
##proportion of each element in a col
probs.bycol = t(count.mat) / col2ncount
##each row: probs for a col
##for each col downsample
##count.down = apply(Hmisc::rMultinom(probs.bycol, col2ncount.down), 1, table) ##Hmisc::rMultinom does not accept different sizes ("m = col2ncount.down").
##rownames(count.down) = rownames(count.mat)
##colnames(count.down) = colnames(count.mat)
count.down = matrix(nrow = nrow(count.mat), ncol = ncol(count.mat), dimnames = list(rownames(count.mat), colnames(count.mat)))
n.cols = length(col2ncount)
for(j.col in 1:n.cols){
jcol.ncount.down = col2ncount.down[j.col]
jcol.probs = probs.bycol[j.col, ]
jcol.count.down = rmultinom(1, jcol.ncount.down, prob = jcol.probs)
count.down[, j.col] = jcol.count.down
}
return(count.down)
}
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