Question: fpkm to tpm conversion
0
gravatar for sayamsmruti
22 months ago by
sayamsmruti0 wrote:

how to convert the fpkm value generated from cufflink to tpm value using r programming????

next-gen R • 3.3k views
ADD COMMENTlink modified 22 months ago • written 22 months ago by sayamsmruti0
1

Actually, you can convert in R by the function I got from another forum and used.

fpkmToTpm <- function(fpkm) {

exp(log(fpkm) - log(sum(fpkm)) + log(1e6))

}

where fpkm is the values you got from TCGA for example.

Luciana

ADD REPLYlink modified 22 months ago • written 22 months ago by lucianalpt20
1

How do you want to cite that in a paper? In general we do not recommend to convert directly between normalized counts because they could been based on whatever non-linear transformation.

ADD REPLYlink modified 22 months ago • written 22 months ago by Michael Dondrup46k

For a small dataset (raw counts) I tested, it did work fine. I did not expect the formula to be so simple :). Thanks for this input. Looking forward to learn more from this discussion.

ADD REPLYlink modified 22 months ago • written 22 months ago by Satyajeet Khare1.4k

Hi

Which package do I need to install for this code?

ADD REPLYlink written 7 months ago by Explorer70
1

Why do you want to use either FPKM or TPM?

Look:

You should abandon RPKM / FPKM. They are not ideal where cross-sample differential expression analysis is your aim; indeed, they render samples incomparable via differential expression analysis:

Please read this: A comprehensive evaluation of normalization methods for Illumina high-throughput RNA sequencing data analysis

The Total Count and RPKM [FPKM] normalization methods, both of which are still widely in use, are ineffective and should be definitively abandoned in the context of differential analysis.

Also, by Harold Pimental: What the FPKM? A review of RNA-Seq expression units

The first thing one should remember is that without between sample normalization (a topic for a later post), NONE of these units are comparable across experiments. This is a result of RNA-Seq being a relative measurement, not an absolute one.

ADD REPLYlink written 7 months ago by Kevin Blighe45k

Please, read this article for basic calculation,

http://bioinfogeek.over-blog.com/2017/09/gene-expression-units-explained-rpm-rpkm-fpkm-and-tpm.html

ADD REPLYlink written 22 months ago by Renesh1.6k

actually after doing cufflink i got the genes.fpkm_tracking as output file so i am clueless what to do next for further data analysis, and how can i convert the generated fpkm values to tpm values...plzz can sum1 help out

ADD REPLYlink written 22 months ago by sayamsmruti0
1

Hi, I highly recommend to leave the cufflinks fpkm output alone and use a more simple and state-of-the-art approach such as featureCounts or HTseq-count directly from BAM files and then generate TPM or CPM from the counts directly using RSEM. In addition I recommend to provide more information, your question is pretty unspecific, and please avoid chat jargon like "plzz sum1". The R-programming portion should be ignored unless there are multiple alternative ways to do this.

ADD REPLYlink modified 22 months ago • written 22 months ago by Michael Dondrup46k
2
gravatar for Satyajeet Khare
22 months ago by
Satyajeet Khare1.4k
Pune, India
Satyajeet Khare1.4k wrote:

It is going to be difficult to calculate TPM from FPKM values in Cuffdiff unless you have raw count values or gene length vector. I would suggest moving to count based methods since the old Tuxedo protocol is deprecated.

ADD COMMENTlink modified 22 months ago • written 22 months ago by Satyajeet Khare1.4k

You can still calculate TPM from RPKM/FPKM values.You need to have information about a total number of transcripts sampled from your read data and avg. a number of nucleotides mapped to each gene.

ADD REPLYlink written 22 months ago by Renesh1.6k

I think the issue is RPKM to TPM conversion in cuffdiff. RPKM values in cuffdiff are internally normalized. When calculated from raw counts, it should not be an issue.

Best

ADD REPLYlink modified 22 months ago • written 22 months ago by Satyajeet Khare1.4k

no, actually it is the fpkm value generated from cufflink, i am unable to convert the fpkm values to tpm

ADD REPLYlink written 22 months ago by sayamsmruti0
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