Entering edit mode

xingxd16
▴
20

@xingxd16-20156
Last seen 4.7 years ago

Hi all

- I use limma to do the differential expression analysis, I want to do the volcano plot, x-axis is logFC and y-axis is
`-1*log10(adjust p)`

. But I found the the adjust p output by limma is so small and after -log10 transfor is up to 300, thats mean the p value gived by limma is`1*E-300`

unit. But in other paper , the adjust p value is about`1*E-30`

unit, and -log10 is about 30. Why the p value is so small in limma? How can I change p value to`1*E-30`

unit. - The logFC is small too. I try to calculate the mean expression for each genes in disease samples and mean value in normal value, then get the logFC, its larger than what limma output . And if I use log2(2) threshold , there is nearly no genes significant express. I have to use log2(1.5) even log2(1.2). Why there are so many genes are so signaficant (with very low p value) but also with a very small logFC at the same times? Can I still consider this genes are differentrial expression(depend on adjust p) even they with small logFC ?

Best

Yes , you are right ! I use limma to do my single cell analysis. The cells in one condition is about 2000+ , the cells in another condition is aboult 3500+ . I just use limma to compare this two conditions follow the "lmFit" and "eBayes" steps .

Yes, that is a huge number of replicates from limma's point of view. It is to be expected that some of the p-values will be very small.

If you use limma for scRNA-seq, it is very important that you only keep genes in the analysis that are detected in a reasonable number of cells. In your case, that might be a few hundred cells.

Good advice ! I should fitler the genes that only express in little percentage of the cells first ! So , you mean that p value and logFC are too samll are reasonable with huge number of replicates ? And can I lower the logFC threshold to identify significant expression genes , such as from log2(2) to log2(1.2) ?

Hi ： I posted another question about limma here , could you help me to answer ! Thanks !