Supplementary: Convert the RNA assay from Assay5 to Assay (MouseBrain_SC)

Seurat objects from version 5 cannot be processed with packageVersion("Seurat") ‘4.3.0’.

We use Seurat ‘5.1.0’ to open the object

# Specify the working directory
setwd(your_working_dir)

# Load necessary packages 
suppressPackageStartupMessages({
  library(hdf5r)
  library(Seurat)
  library(dplyr)
  library(RColorBrewer)
  library(CellTrek)
  library(ggplot2)
  library(spacexr)
  library(paletteer)
})
# Check Seurat version
packageVersion("Seurat")

# Load your Seurat v5 object
V5_Path <- 'MouseBrain_snRNA_V5_obj.rds'

Brain_SC <- readRDS(V5_Path)
Brain_SC$Annotation = Idents(Brain_SC)
Brain_SC
head(Brain_SC@meta.data)

# Normalization and runPCA
DefaultAssay(Brain_SC) = 'RNA'
Brain_SC <- NormalizeData(Brain_SC, assay='RNA') %>% 
  FindVariableFeatures() %>% 
  ScaleData(features=rownames(Brain_SC))
Brain_SC <- RunPCA(Brain_SC, features=VariableFeatures(Brain_SC),
                         verbose = F)

ElbowPlot(Brain_SC, ndims = 50) 

# RunUMAP
Brain_SC <- FindNeighbors(Brain_SC, reduction='pca', dims = 1:25)
Brain_SC <- FindClusters(Brain_SC, resolution = 0.1)
Brain_SC <- RunUMAP(Brain_SC, dims = 1:25)
Idents(Brain_SC) <- Brain_SC$Annotation
DimPlot(Brain_SC)

# Convert the RNA assay from Assay5 to Assay
Brain_SC[["RNA"]] <- as(object = Brain_SC[["RNA"]], Class = "Assay")

# Update the Seurat object to ensure it matches the Seurat v4 format
Brain_SC <- UpdateSeuratObject(Brain_SC)

# Save the updated Seurat object
saveRDS(Brain_SC, file = "MouseBrain_snRNA_V4_obj.rds")

# Print the new Seurat object to verify
print(Brain_SC)