###### tags: `ノート` `TS` `解析` `single-cell RNA-seq`

# 22-03： ES由来セルトリ細胞scRNAseq解析　その３　各マーカーのplot (aggrしない)

<br>

#### 2026/04/22
#### 【 作業場所＆保存場所 】

• 作業場所
　~/osc-fs/22_TS_FTSLC_scRNAseq_2022/Seurat_Non_aggr

---

#### ※ aggrしないでSeuratした方

【参考】
scRNA-seqデータの解析(応用編)
https://hackmd.io/@takahirosuzuki/ByoEBNVPL
<br>

### 1. 各マーカーのfeatureplot

Early-Sertoli: Sry, Runx1, Nr0b1
Sertoli: Sox9, Dhh, Amh
Early Prog/Int. Prog: Nr2f1, Tcf21, Pdgfra
Getal Leydig:Cyp11a1, Cyp17a1, Star
の発現をfeatureplot, 各サンプル毎で。

load("20220422_5_data.integrated.RData")

#Early-Sertoli

pdf("FeaturePlot_Sry.pdf", width=35, height=5)
FeaturePlot(object=data.integrated, features= "Sry", col=c("gray", "red"), split.by="sample", min.cutoff=0.1, pt.size=1)
dev.off()

pdf("FeaturePlot_Runx1.pdf", width=35, height=5)
FeaturePlot(object=data.integrated, features= "Runx1", col=c("gray", "red"), split.by="sample", min.cutoff=0.1, pt.size=1)
dev.off()

pdf("FeaturePlot_Nr0b1.pdf", width=35, height=5)
FeaturePlot(object=data.integrated, features= "Nr0b1", col=c("gray", "red"), split.by="sample", min.cutoff=0.1, pt.size=1)
dev.off()

pdfjoin --outfile FeaturePlot_Early-Sertoli.pdf FeaturePlot_Sry.pdf FeaturePlot_Runx1.pdf FeaturePlot_Nr0b1.pdf


#Sertoli

pdf("FeaturePlot_Sox9.pdf", width=35, height=5)
FeaturePlot(object=data.integrated, features= "Sox9", col=c("gray", "red"), split.by="sample", min.cutoff=0.1, pt.size=0.5)
dev.off()

pdf("FeaturePlot_Dhh.pdf", width=35, height=5)
FeaturePlot(object=data.integrated, features= "Dhh", col=c("gray", "red"), split.by="sample", min.cutoff=0.1, pt.size=0.5)
dev.off()

pdf("FeaturePlot_Amh.pdf", width=35, height=5)
FeaturePlot(object=data.integrated, features= "Amh", col=c("gray", "red"), split.by="sample", min.cutoff=0.1, pt.size=0.5)
dev.off()

pdfjoin --outfile FeaturePlot_Sertoli_0.5.pdf FeaturePlot_Sox9.pdf FeaturePlot_Dhh.pdf FeaturePlot_Amh.pdf


#Early Prog/Int. Prog: Nr2f1, Tcf21, Pdgfra

pdf("FeaturePlot_Nr2f1.pdf", width=35, height=5)
FeaturePlot(object=data.integrated, features= "Nr2f1", col=c("gray", "red"), split.by="sample", min.cutoff=0.1, pt.size=1)
dev.off()

pdf("FeaturePlot_Tcf21.pdf", width=35, height=5)
FeaturePlot(object=data.integrated, features= "Tcf21", col=c("gray", "red"), split.by="sample", min.cutoff=0.1, pt.size=1)
dev.off()

pdf("FeaturePlot_Pdgfra.pdf", width=35, height=5)
FeaturePlot(object=data.integrated, features= "Pdgfra", col=c("gray", "red"), split.by="sample", min.cutoff=0.1, pt.size=1)
dev.off()

pdfjoin --outfile FeaturePlot_EarlyProgInt.pdf FeaturePlot_Nr2f1.pdf FeaturePlot_Tcf21.pdf FeaturePlot_Pdgfra.pdf


#Getal Leydig:Cyp11a1, Cyp17a1, Star

pdf("FeaturePlot_Cyp11a1.pdf", width=35, height=5)
FeaturePlot(object=data.integrated, features= "Cyp11a1", col=c("gray", "red"), split.by="sample", min.cutoff=0.1, pt.size=1)
dev.off()

pdf("FeaturePlot_Cyp17a1.pdf", width=35, height=5)
FeaturePlot(object=data.integrated, features= "Cyp17a1", col=c("gray", "red"), split.by="sample", min.cutoff=0.1, pt.size=1)
dev.off()

pdf("FeaturePlot_Star.pdf", width=35, height=5)
FeaturePlot(object=data.integrated, features= "Star", col=c("gray", "red"), split.by="sample", min.cutoff=0.1, pt.size=1)
dev.off()

pdfjoin --outfile FeaturePlot_GetalLeydig.pdf FeaturePlot_Cyp11a1.pdf FeaturePlot_Cyp17a1.pdf FeaturePlot_Star.pdf


#overy-specific marker: Foxl2、Wnt4、Dax１

pdf("FeaturePlot_Foxl2.pdf", width=35, height=5)
FeaturePlot(object=data.integrated, features= "Foxl2", col=c("gray", "red"), split.by="sample", min.cutoff=0.1, pt.size=0.5)
dev.off()

pdf("FeaturePlot_Wnt4.pdf", width=35, height=5)
FeaturePlot(object=data.integrated, features= "Wnt4", col=c("gray", "red"), split.by="sample", min.cutoff=0.1, pt.size=0.5)
dev.off()

pdf("FeaturePlot_Nr0b1.pdf", width=35, height=5)
FeaturePlot(object=data.integrated, features= "Nr0b1", col=c("gray", "red"), split.by="sample", min.cutoff=0.1, pt.size=0.5)
dev.off()

pdfjoin --outfile FeaturePlot_GetalLeydig.pdf FeaturePlot_Cyp11a1.pdf FeaturePlot_Cyp17a1.pdf FeaturePlot_Star.pdf


あと、Sryですが、XYのE11.5で少し発現している細胞がありそうなのですが、pt.sizeやmax.cutoffあたりをいじって少し見やすくできますか？

#FeaturePlot_Sry.pdf
FeaturePlot(object=data.integrated, features= "Sry", col=c("gray", "red"), split.by="sample", min.cutoff=0.1, pt.size=0.5)
FeaturePlot(object=data.integrated, features= "Sry", col=c("gray", "red"), split.by="sample", min.cutoff=0.5, pt.size=0.5)


<br>

#### 2. まとめ　MultiVlnPlot

multiVlnPlot <- function(object, features, pt.size = 0, log = FALSE, title = NULL, out_file = NULL, ...){
    library(Seurat)
    library(ggplot2)
    library(cowplot)

    scaleFUN <- function(x) sprintf("%.1f", x)

    gplot.list <- list()
    for(i in 1:length(features)){
        vg <- VlnPlot(object = object, features = features[i], pt.size = pt.size, log = log,...)
        vg <- vg + scale_y_continuous(labels=scaleFUN)
        vg <- vg + scale_fill_discrete(name="Cluster")
        vg <- vg + ylab(features[i])
        vg <- vg + theme(legend.position = 'none',
                         axis.title.x = element_blank(),
                         axis.title.y = element_text(face="bold", vjust = 0.5),
                         plot.title = element_blank(),
                         axis.text.x = element_blank(),
                         panel.border = element_rect(colour="gray1", fill=NA)
                        )
        gplot.list <- c(gplot.list, list(vg))
    }

    legend <- get_legend(gplot.list[[1]] + 
                        theme(legend.position = "right") +
                        guides(color = guide_legend(ncol = 1))
    )

    # now add the title
    if(!is.null(title)){
        gtitle <- ggdraw() + 
            draw_label(
                title,
                fontface = 'bold',
                x = 0.5,
                hjust = 0.5
            ) 
    }else gtitle <- NULL

    gplot_all <- plot_grid(plotlist = gplot.list, ncol=1)
    gplot_all_title <- plot_grid(gtitle, 
                        gplot_all, nrow = 2, 
                        rel_heights = c(0.5/(length(features)+0.5), length(features)/(length(features)+0.5))
                        )

    gplot_all_legend <- plot_grid(gplot_all_title, legend, ncol=2, rel_widths = c(8, .6))

    plot(gplot_all_legend)

    if (!is.null(out_file)){
        ggsave(file=out_file, plot=gplot_all_legend, width=14, height=length(features)+0.5)
    }
    return (gplot_all_legend)
}


#マーカーの設定
markers_Sertoli <- c("Sox9", "Dhh", "Amh")

# markers_SertoliでPlot
multiVlnPlot(object=data.integrated, features = markers_Sertoli, title="Sertoli : Sox9, Dhh, Amh Markers", out_file = "MultiVlnPlot_Sertoli_markers.pdf")