Modeling expression ranks for noise-tolerant differential expression analysis of scRNA-Seq data
ROSeq - A rank based approach to modeling gene expression with filtered and normalized read count matrix. ROSeq takes filtered and normalized read matrix and cell-annotation/condition as input and determines the differentially expressed genes between the contrasting groups of single cells. One of the input parameters is the number of cores to be used.
The developer’s version of the R package can be installed with the following R commands:
if (!requireNamespace("BiocManager", quietly = TRUE)) install.packages("BiocManager") # The following initializes usage of Bioc devel BiocManager::install(version='devel') BiocManager::install("ROSeq")
The github’s version of the R package can be installed with the following R commands:
This vignette uses the Tung dataset, which is already inbuilt in the package, to demonstrate a standard pipeline.
Libraries need to be loaded before running.
samples<-list() samples$count<-ROSeq::L_Tung_single$NA19098_NA19101_count samples$group<-ROSeq::L_Tung_single$NA19098_NA19101_group samples$count[1:5,1:5] #> NA19098.r1.A01 NA19098.r1.A02 NA19098.r1.A03 NA19098.r1.A04 #> ENSG00000237683 0 0 0 1 #> ENSG00000187634 0 0 0 0 #> ENSG00000188976 3 6 1 3 #> ENSG00000187961 0 0 0 0 #> ENSG00000187583 0 0 0 0 #> NA19098.r1.A05 #> ENSG00000237683 0 #> ENSG00000187634 0 #> ENSG00000188976 4 #> ENSG00000187961 0 #> ENSG00000187583 0
Below commands can be used for Cell/gene filtering, TMM normalization and voom transformation. The user is free to use an alternative preprocessing strategy while using different filtering/normalization methods.
gene_names<-rownames(samples$count) samples$count<-apply(samples$count,2,function(x) as.numeric(x)) rownames(samples$count)<-gene_names samples$count<-samples$count[,colSums(samples$count> 0) > 2000] gkeep<-apply(samples$count,1,function(x) sum(x>2)>=3) samples$count<-samples$count[gkeep,] samples$count<-limma::voom(ROSeq::TMMnormalization(samples$count))
Input: gene expression matrix with genes in rows and cells in columns. Condition/group annotation of cells also need to be supplied. User can set numCores based the hardware specifications in her computer.