How to perform a methylation analyisis from m values
Entering edit mode
Irene • 0
Last seen 2 days ago

I am trying to perform a methylation analysis from a txt file which contains the m values.
The RNA samples were taken to a company that performed an Illumina EPIC array.
They performed the first steps of the analysis which consisted of the pre-processing and quality control of the data using minfi package.
Then, they sent us two txt files one with the m-values and the other with the b-values.
The structure of these data in the txt file is (I am showing only the first 2 lines of17.000)
island patient1 patient2 patient3 control1 control2 control3
cg14817997 3.09546 2.11941 2.90160 1.91566 2.45522 2.36514

My doubt is when it comes to start the analysis from these data. I understand that I have to use the function:

makeGenomicRatioSetFromMatrix(mat, rownames = NULL, pData = NULL,
                              array = "IlluminaHumanMethylation450k",
                              annotation = .default.450k.annotation,
                              mergeManifest = FALSE, what = c("Beta", "M"))

It creates a GenomicRatioSet object that stores the methylation data along with additional information such as genomic coordinates of regions, genomic annotations and sample meta-information.
This function requires a matrix and I don't know how to set it up. I also think that I need to say R which are my groups so I create this:

df <- data.frame( test=1, d=c(rep('patient', 3 ), rep('control', 3)) )
df$d <- factor( df$d, levels = c('patient', 'control'))

However, I need help constructing the matrix correctly so that the function makeGenomicRatioSetFromMatrix works correctly and can search for differentially methylated positions.

Any help is welcome.

Thank u

MethylationArrayData methylationArrayAnalysis • 288 views
Entering edit mode
Last seen 1 day ago
United States

You could create the matrix by doing something like this.

mat <- as.matrix(read.table(<filename goes here>, header = TRUE, row.names = 1, sep = "\t"))
## you might need to adjust the 'sep' argument, depending on the separator for your text file.
rgset <- makeGenomicRatioSetFromMatrix(mat, pData = df, 
                                       array = "IlluminaHumanMethylationEPIC",
                                       annotation = "IlluminaHumanMethylationEPICanno.ilm10b4.hg19")

That's untested code, as I don't have your text file, but it should get you close.

Entering edit mode

Thank you very much @JamesW.MacDonald, it worked!

Now I need to obtain the different methylation positions (DMPs)

I think that the function needed is "dmpFinder":

dmpFinder(dat, pheno, type = c("categorical", "continuous"),
    qCutoff = 1, shrinkVar = FALSE)

However, I don't know if I am using the function correctly. In my case, I substitute the arguments in the following way, but when I run the function, I get the following error

result<-dmpFinder(mat, df$d, type = c("categorical"),
    qCutoff = 1, shrinkVar = FALSE)

Error in dmpFinder(mat, df$d, type = c("categorical"), qCutoff = 1, shrinkVar = FALSE) : 
  is.numeric(dat) is not TRUE

Maybe I'm not using the right function either...

Thank u


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