inferCNV鉴定肿瘤中的恶性细胞流程

1、准备相关文件

sce <- readRDS("path_to_seurat/seurat.rds")
#鉴定内皮细胞中的恶性细胞
epi.cells  <- row.names(sce@meta.data)[which(sce$free_annotation=='Epithelial cells')]
epi_sce <- subset(sce, cells = epi.cells)
epi_sce@meta.data
length(epi.cells)
#Raw Counts Matrix for Genes x Cells
epiMat=as.data.frame(GetAssayData(subset(sce, cells=epi.cells)))
#细胞的注释文件 共两列，一列CB，一列细胞来源，tab分割，无列名
groupinfo=data.frame(v1=colnames(epiMat),
                     v2=epi_sce@meta.data$type)
#基因排序文件
geneInfor=annoGene(rownames(epiMat),"SYMBOL",'human')
colnames(geneInfor)
geneInfor=geneInfor[with(geneInfor, order(chr, start)),c(1,4:6)]
geneInfor=geneInfor[!duplicated(geneInfor[,1]),]
length(unique(geneInfor[,1]))
head(geneInfor)

## 这里可以去除性染色体
# 也可以把染色体排序方式改变
dat=epiMat[rownames(epiMat) %in% geneInfor[,1],]
dat=dat[match( geneInfor[,1], rownames(dat) ),] 
dim(dat)
expFile='path_to_save/expFile.txt'
write.table(dat,file = expFile,sep = '\t',quote = F)
groupFiles='path_to_save/groupFiles.txt'
head(groupinfo)
write.table(groupinfo,file = groupFiles,sep = '\t',quote = F,col.names = F,row.names = F)
head(geneInfor)
geneFile='path_to_save/geneFile.txt'
write.table(geneInfor,file = geneFile,sep = '\t',quote = F,col.names = F,row.names = F)

2、运行inferCNV

infercnv_obj = CreateInfercnvObject(raw_counts_matrix=expFile,
                                    annotations_file=groupFiles,
                                    delim="\t",
                                    gene_order_file= geneFile,
                                    ref_group_names=c("normal"))  ## 这个取决于自己的分组信息里面的
# 手动指定输出目录
out_dir <- "path_to_save_inferCNV_result"
infercnv_obj = infercnv::run(infercnv_obj,
                             cutoff=0.1, # cutoff=1 works well for Smart-seq2, and cutoff=0.1 works well for 10x Genomics
                             out_dir=out_dir, 
                             cluster_by_groups=TRUE, 
                             denoise=TRUE,
                             HMM=TRUE)

3、提取恶性细胞

# 读取预测的CNV区域和基因文件
cnv_regions <- read.table("path_to_save_inferCNV_result/17_HMM_predHMMi6.hmm_mode-samples.pred_cnv_regions.dat", header = TRUE)
cnv_genes <- read.table("path_to_save_inferCNV_result/17_HMM_predHMMi6.hmm_mode-samples.pred_cnv_genes.dat", header = TRUE)

# 读取细胞分组信息
cell_groupings <- read.table("path_to_save_inferCNV_result/17_HMM_predHMMi6.hmm_mode-samples.cell_groupings", header = TRUE)

# 识别具有显著CNV的细胞
malignant_cells <- cell_groupings[cell_groupings$cell_group_name %in% cnv_regions$cell_group, ]

# 查看恶性细胞
malignant_cells_need <- malignant_cells$cell
malignant_cells_need