---
title: "prepare EGFP markers"
author: "Yifang Liu"
date: "`r Sys.Date()`"
output:
  rmdformats::html_clean:
    code_folding: hide
    fig_width: 10
    fig_height: 10
    highlight: kate
    thumbnails: false
    lightbox: true
    gallery: true
---

```{r knitr_init, echo=FALSE, cache=FALSE}
library(knitr)
library(rmdformats)

options(max.print = 200)
opts_chunk$set(echo = TRUE,
               cache = FALSE,
               prompt = FALSE,
               tidy = TRUE,
               comment = NA,
               message = FALSE,
               warning = FALSE,
               dev = c('png', 'pdf'),
               fig.width = 10,
               fig.height = 10,
               fig.align = "center",
               fig.path = 'PDF/',
               dpi = 72)
opts_knit$set(width = 75)
```

```{r setup}
set.seed(123)

npc <- 20
# theta1 <- 2
# theta2 <- 5
# theta <- c(theta1, theta2)
resolution <- 0.1
pt_size <- 2

# Suppress loading messages
suppressPackageStartupMessages({
  library(Matrix)
  library(dplyr)
  library(tidyverse)
  library(Seurat)
  library(cowplot)
  library(Rcpp)
  library(harmony)
  library(SoupX)
})
```

```{r load_data}
EGFP <- readRDS("Data/2020-02-05_EGFP_seurat_obj.Rds")
DefaultAssay(EGFP) <- "RNA"
selected_res <- paste0("RNA_snn_res.", resolution)
Idents(EGFP) <- selected_res

table_df <- table(EGFP@meta.data[ , selected_res], EGFP@meta.data$LibraryID) %>%
    as.data.frame() %>% spread(key = Var2, value = Freq)
```

```{r markers}
if (file.exists("Markers/2020-02-05_EGFP_markers.Rds")) {
    markers_list <- readRDS("Markers/2020-02-05_EGFP_markers.Rds")
} else {
  dir.create("Markers")
  table_df <- table(EGFP@meta.data[ , selected_res], EGFP@meta.data$LibraryID) %>% as.data.frame() %>% spread(key = Var2, value = Freq)
  colnames(table_df)[1] <- "cluster"
  # str(table_df)
  markers_list <- list()
  for(cluster_id in table_df$cluster){
      markers <- FindMarkers(EGFP, ident.1 = cluster_id)
      # markers$gene <- row.names(markers)
      write.csv(markers, file = paste0("Markers/2020-02-05_EGFP_markers_cluster", cluster_id, ".csv"))
      markers_list[[paste0("cluster", cluster_id)]] <- markers
  }
  saveRDS(markers_list,
          file = "Markers/2020-02-05_EGFP_markers.Rds")
}
```

# Notes

2020-02-05:

  * only EGFP.

Sat Nov 30, 2019:

  * prepare markers.

Tue Oct 29, 2019:

  * use SoupX fixed 0.45 to remove ambient RNA.

Mon Oct 7, 2019:

  * Add more sequence depth.

Mon, Sep 30, 2019:

  * remove genes: EGFP, Tsc1, gig. Then perform integrate analysis of EGFP, TSC1.

Fri, Sep 20, 2019:

  * First version for integrate analysis of EGFP, TSC1.

# Session Info

```{r sessioninfo, message=TRUE}
sessionInfo()
```