When I tried to read in the sample sheet (txt file) as a table using DESeqDataSetFromHTSeqCount, I got an error message: Error in file(file, "rt") : cannot open the connection.

# Read in sample sheet and append sample name as row labels
# Args:
#   input.file: a .txt samplesheet
#
# Returns:
#   table: a data table

ReadTable <- function(input.file) {
  table <- read.table(input.file, header=T, sep="\t")
  rownames(table) <- table$SampleName
  return(table)
}


sampleTable <- ReadTable("sample_sheet_copy.txt")
sampletable_med <- subset(sampleTable, Tissue == "med")
sampletable_cerv <- subset(sampleTable, Tissue == "cerv")

# load in the count data to produce a count matrix
# directory is path to directory where the counts are stored (one per sample)
# design is how we wish to model the data: here we are measuring differences by genotype

ddsHTSeq <- DESeqDataSetFromHTSeqCount(sampleTable = sampleTable,
                                       directory = "counts_4thcol",
                                       design = ~ 1)

counts_4thcol is a folder containing reads per gene in out.tab format, for example:

Xkr4 944

Gm39585 1

Gm38717 2

Rp1 0

Sox17 338 ...

Sample sheet is in a txt file like the following:

SampleName FileName Genotype Age Tissue

E1061-med_S4_merged E1061-med_S4_merged_counts.txt mdx 12m med ...

R is telling you that the file doesn't exist in your current working directory. Here's an example:

> read.table("this_file_does_not_exist.txt")
Error in file(file, "rt") : cannot open the connection
In addition: Warning message:
In file(file, "rt") :
  cannot open file 'this_file_does_not_exist.txt': No such file or directory

So you either mis-typed the file name, are in the wrong working directory, or the file just doesn't exist.