not centered volcano plot, Seurat

Question

[sooni]

Hello.

I performed DE analysis on single cell data using Seurat, following the pseudobulk tutorial on this page. After DE analysis, I drew a volcano plot, and the result is a non-centered volcano plot.

The center of the volcano plot is not located at 0 on the x-axis. I looked into the reason for this and found that this problem occurs when there is a problem with fc.slot when analyzing using the FindMarkers() function in Seurat, but I couldn't find a proper solution. How can I solve this problem? I used Seurat v.5.

Please help me. Thank you for help!

[ATpoint]

My suggestions would be to plot an MA-plot first, and show this. Usually non-centered plots indicate poor normalization. Which test did you run?

[sooni]

I ran DESeq2 as a test! Bottom line is, I solved this problem! It was caused by creating pseudo-bulk data and not running the NormalizeData() function. This problem is now solved.