I want to do DE analysis using DESeq2. My experiment is small RNAseq experiment with 5 tissue samples. I want to find out the DE genes between different tissues.
I have following matrix (as an example)
My aim is to find DE gene between each column, - i.e. tissue1 vs tissue2, tissue1 vs tissue3 ......tissue2 vs tissue3...... tissue4 vs tisue5
I dont fully understand the program, i tried following the program vignette as follows:
> CountTable = read.table("test.tsv", header=TRUE, row.names=1)
gene tissue1 tissue2 tissue3 tissue4
gene1 233 91 17 593
gene2 1011 0 7 1
gene3 963 2 3 66
gene4 908 41 1 74
gene5 596 50 26 328
gene6 1 0 0 0
> colData = data.frame(
+ row.names= colnames(CountTable),
+ condition = c("tissue1", "tissue2", "tissue3", "tissue4", "tissue5"),
+ libType = c( "single-end", "single-end", "single-end", "single-end", "single-end"))
> dds <- DESeqDataSetFromMatrix( countData = CountTable, colData = colData,
> design = ~ condition)
> dds <- DESeq(dds)
#estimating size factors
#gene-wise dispersion estimates
#final dispersion estimates
#fitting model and testing
#In checkForExperimentalReplicates(object, modelMatrix) :
#same number of samples and coefficients to fit,
#estimating dispersion by treating samples as replicates.
#read the ?DESeq section on 'Experiments without replicates'
>res <- results(dds)
#log2 fold change (MAP): condition testis vs brain
#Wald test p-value: condition tissue5 vs tissue1
#DataFrame with 927 rows and 6 columns
baseMean log2FoldChange lfcSE stat pvalue padj
So in the result I am getting comparison only between tissue5 and tissue1, what do i need to do to find out comparison between each tissue?
Help is greatly appreciated.
P.S. : I am new to R and first time using DESeq2