代码库3-Signac代码

library(Signac)
counts <- Read10X_h5("../vignette_data/atac_v1_pbmc_10k_filtered_peak_bc_matrix.h5")
fragment_file <- '../vignette_data/atac_v1_pbmc_10k_fragments.tsv.gz'
chrom_assay <- CreateChromatinAssay(counts,sep = c(":", "-"),fragments = fragment_file)
SeuratObj_ATAC <- CreateSeuratObject(chrom_assay,assay = "ATAC")
# SeuratObj_ATAC <- CreateSeuratObject(counts, assay = "ATAC")

SeuratObj_ATAC <- RunTFIDF(SeuratObj_ATAC)
SeuratObj_ATAC <- FindTopFeatures(SeuratObj_ATAC, min.cutoff = 'q0')
SeuratObj_ATAC <- RunSVD(SeuratObj_ATAC)
DepthCor(SeuratObj_ATAC)
SeuratObj_ATAC <- RunUMAP(SeuratObj_ATAC, dims = 2:50, reduction = 'lsi', 
                     reduction.name = 'atac_umap', reduction.key = 'atac_umap')

SeuratObj_ATAC <- FindNeighbors(object = SeuratObj_ATAC, reduction = 'lsi', dims = 2:30)
SeuratObj_ATAC <- FindClusters(object = SeuratObj_ATAC, verbose = FALSE, algorithm = 3)
DimPlot(object = SeuratObj_ATAC, label = TRUE) + NoLegend()

# GeneActivity
library(EnsDb.Hsapiens.v75)
annotations <- GetGRangesFromEnsDb(ensdb = EnsDb.Hsapiens.v75)
seqlevelsStyle(annotations) <- 'UCSC'
genome(annotations) <- "hg19"
Annotation(SeuratObj_ATAC) <- annotations
gene.activities <- GeneActivity(SeuratObj_ATAC)
SeuratObj_ATAC[['ACTIVITY']] <- CreateAssayObject(counts = gene.activities)
SeuratObj_ATAC <- NormalizeData(SeuratObj_ATAC,assay = 'ACTIVITY')
DefaultAssay(SeuratObj_ATAC) <- 'ACTIVITY'
SeuratObj_ATAC <- FindVariableFeatures(SeuratObj_ATAC, selection.method = "vst", nfeatures = 2000)          
LabelPoints(plot = VariableFeaturePlot(SeuratObj_ATAC), points = head(VariableFeatures(SeuratObj_ATAC), 10), repel = TRUE)
SeuratObj_ATAC <- ScaleData(SeuratObj_ATAC)    
SeuratObj_ATAC <- RunPCA(SeuratObj_ATAC)   
ElbowPlot(SeuratObj_ATAC)
SeuratObj_ATAC <- RunUMAP(SeuratObj_ATAC, dims = 1:20, reduction.name = 'ga_umap', reduction.key = 'ga_umap')   
DimPlot(SeuratObj_ATAC, reduction = "ga_umap",group.by = 'true',label = T) + NoLegend()

Idents(SeuratObj_ATAC) <- 'true'
CoveragePlot(SeuratObj_ATAC,region = c('chr5-54070562-54071062','chr1-212806552-212807052'),
             extend.upstream = 40000,extend.downstream = 20000)

参考

https://stuartlab.org/signac/articles/pbmc_vignette

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