Is there a way to calculate p-values, FDR or t test starting from logFC data?
3
2
Entering edit mode
7.3 years ago
Spacebio ▴ 200

I am new in DEA and I have RNA-Seq data already normalised to TPM, I calculated the log2FC values but I would like to calculate the p-values, FDR or t-values. Is there a way to do this kind of analysis in R? Here people discuss about it but the data is normalised in a different way and they didn't really answer the question. Any help is very appreciated.

R TPM • 4.7k views
ADD COMMENT
1
Entering edit mode

Hi Spacebio,

Do not make redundant posts with similar question. I found this thread started by you TPM to logFC and pvalues

ADD REPLY
0
Entering edit mode

Hi EagleEye, I'm sorry! I am learning to use this platform to ask questions or debate about some topics. I couldn't find my old question, for that reason I started a new one. Please be a bit patient, slowly but surely I am learning to handle the info here.

ADD REPLY
2
Entering edit mode
7.3 years ago
Spacebio ▴ 200

I found this question so I guess that is a helpful way to do so.

d <- DGEList(counts = myDF[,2:7], group = c(rep("S1", 3), rep("S2", 3)), genes = myDF[,1])
d <- calcNormFactors(d)
d <- estimateCommonDisp(d)
d <- estimateTagwiseDisp(d)
d <- estimateTrendedDisp(d)
de <- exactTest(d, pair=c("S1", "S2"), dispersion = "tagwise")
deDF <- as.data.frame(de)

Any other ideas? I heard there is a way to convert TPM to raw counts, is that true? Any help is very appreciated!

ADD COMMENT
0
0
Entering edit mode

Hello, thank you for your help and suggestions! The problem here is that I have a .csv file containing RNA-Seq values normalised already to TPM, I have no access to the raw data. That is the reason why I am performing the analysis by hand. I calculated the log2FC values but now I would like to calculate the p-values, FDR or t-test in order to make sure that my results make sense. I am using R, but most of the packages ask for the raw data. Any other suggestion?

ADD REPLY
0
Entering edit mode

you don't need raw count values, you can use TPM and limma to get DEGs

ADD REPLY
0
Entering edit mode
7.3 years ago
theobroma22 ★ 1.2k

You can use different R packages like QValue available in Bioconductor, or R functions like the p-value and t-test function. Their usages are pretty straightforward, but if you have issues just start a new post so we can try to help you.

ADD COMMENT
0
Entering edit mode

Hello, thanks for your suggestion. The problem here is that R packages ask for the raw data and I have no access to it. My data is a .csv file containing RNA-SEQ data normalised already to TPM.

ADD REPLY
0
Entering edit mode

Typically after normalization comes differential expression analysis. This is done by developing a model, linear or other. So, since you have normalized values now you can start to compare your null and full models. From this comparison you can get your p-values, q-values and FDR. In summary, you don't have to start with the raw data to finish / complete your analysis.

ADD REPLY
0
Entering edit mode

I didn't know that! Thank you very much for the info, I am new in this kind of analysis and there is a lot of info I am missing.

ADD REPLY

Login before adding your answer.

Traffic: 1595 users visited in the last hour
Help About
FAQ
Access RSS
API
Stats

Use of this site constitutes acceptance of our User Agreement and Privacy Policy.

Powered by the version 2.3.6