How do we identify conserved cell type markers after SCTransform - IntegrateLayers in Seurat 5? #8740
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greengeckoruns
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I also would like some clarification on how SCTransform should be used with the integrative analysis pipeline. |
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Hello, and thanks to everyone in advance for reading this question. I use Seurat 5 to analyze a single-cell experiment with two conditions (A vs. B). At the moment, I am trying out different data (pre)processing steps (NormalizeData-FindVariableFeatures-ScaleData [NFS] vs. SCTransform [SCT]) and integrations (cca, rpca, fastnmm, harmony, sci) to learn how different approaches influence the interpretation of results. My starting Seurat object has 2 layers (counts.A and counts.B), and the code for the NFS processing pipeline looks something like this:
The above works nicely irrespective of the integration method used (cca, rpca, fastnmm, harmony, scvi) during the IntegrateLayers() function call. However, I am having issues using FindConservedMarkers() on the SCT processed Seurat object. The code is the following:
However, when I now try to run:
FindConservedMarkers(obj_SCT, ident.1 = 1, grouping.var = "orig.ident")
I get the following error, irrespective of whether the PrepSCTFindMarkers(obj_SCT) has been called or not:
On the other hand,
FindMarkers(obj_SCT, ident.1 = 1, ident.2 = 5)
runs as expected after calling PrepSCTFindMarkers(obj_SCT).I would appreciate any suggestions on what I am doing wrong. I also apologize if I am missing something obvious, and thanks for reading this post/helping.
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