harmony与sct与harmony seurat中的 SCT连用

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已于 2024-08-22 22:00:52 修改

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分类专栏：纸上得来终觉浅文章标签： linux 运维服务器

于 2023-03-06 16:37:28 首次发布

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该文展示了如何利用Seurat包中的SCTransform对单细胞测序数据进行预处理，然后通过Harmony方法校正样本间的批次效应。PCA、UMAP和TSNE被用于降维和可视化，以揭示细胞间的相互关系和潜在的细胞类型结构。

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Harmony with Seurat SCTransform · Issue #41 · immunogenomics/harmony (github.com)https://github.com/immunogenomics/harmony/issues/41

pbmc_harmony_integrated <- merge(pbmc1, 
y=c(pbmc2,pbmc3, pbmc4), 
add.cell.ids = c("Donor1","Donor2","Donor3","Donor4"), 
project = "pbmc_combined") %>% 

SCTransform(vars.to.regress = "percent.mt",
 return.only.var.genes =FALSE, verbose = TRUE) %>% 

 RunPCA(pc.genes = pbmc_integrated@var.genes, verbose = FALSE) %>% 
RunHarmony(c("donor.number"),plot_convergence = TRUE)

SCTransform默认作用于RNA assay

SCTransform(
  object,
  assay = "RNA",
  new.assay.name = "SCT",
  reference.SCT.model = NULL,
  do.correct.umi = TRUE,
  ncells = 5000,
  residual.features = NULL,
  variable.features.n = 3000,
  variable.features.rv.th = 1.3,
  vars.to.regress = NULL,
  do.scale = FALSE,
  do.center = TRUE,
  clip.range = c(-sqrt(x = ncol(x = object[[assay]])/30), sqrt(x = ncol(x =
    object[[assay]])/30)),
  conserve.memory = FALSE,
  return.only.var.genes = TRUE,
  seed.use = 1448145,
  verbose = TRUE,
  ...
)

.libPaths(c("/home/data/refdir/Rlib/",  "/home/data/t040413/R/x86_64-pc-linux-gnu-library/4.2", "/usr/local/lib/R/library"))
library(Seurat)
library(dplyr)
library(tibble)
library(ggplot2)
library(dplyr)
library(tibble)
library(stringr)

load("./GSE183852_DCM_Integrated.Robj")




#DefaultAssay(RefMerge)="prediction.score.celltype.l2"
RefMerge$stim=str_replace_all(colnames(RefMerge),pattern ="[A|T|C|G]",replacement = "")
Idents(RefMerge)=RefMerge$stim
table(RefMerge$stim)


DefaultAssay(RefMerge)
table(RefMerge$Names)
table(RefMerge$tech)


Idents(RefMerge)=RefMerge$Names
Idents(RefMerge)=RefMerge$tech
All.merge=subset(RefMerge,idents = "SN")

DefaultAssay(All.merge)="SCT"
head(All.merge@meta.data)

#################################
All.merge=SCTransform(All.merge, verbose = FALSE)  %>% RunPCA(npcs = 50, verbose = FALSE)
library('harmony')
All.merge <- All.merge  %>% RunHarmony("stim", plot_convergence = TRUE)
harmony_embeddings <- Embeddings(All.merge , 'harmony') 
#######################cluster
dims = 1:30
All.merge  <- All.merge  %>% 
  RunUMAP(reduction = "harmony", dims = dims) %>% 
  RunTSNE(reduction = "harmony", dims = dims) %>% 
  FindNeighbors(reduction = "harmony", dims = dims)


save(All.merge , file="All.merge .rds")
getwd()
list.files()

setwd("D:/yll/heart_fibroblast/project/gse183852")