Compact transcription factor cassettes generate functional, engraftable motor neurons by direct conversion

Pre-publication release:

Item	DOI
Article
Dataset for python
scRNAseq data I
scRNAseq data II

For python analysis

Setting data directory

We use rushd to organge our data analysis. rushd requires you to specify a data directory. After you decide where to put the data locally, create a file called datadir.txt in the GitHub repo that contains the code. The datadir.txt file will contain only the absolute path to the folder in which you placed the data.

For example, if you placed the GitHub repo in /Users/username/Documents/GitHub/article-engraftable-neurons/ and you place the data in /Users/username/Documents/GitHub/article-engraftable-neurons/data/, then you would place the datadir.txt file in /Users/username/Documents/GitHub/article-engraftable-neurons/ and it should contain one line:

/Users/username/Documents/GitHub/article-engraftable-neurons/data/

Instructions

1. Download local copy of data
2. Update datadir.txt to point to data folder
3. Use virtual environment with Python 3.9.6
4. Install packages

pip install -r requirements.txt

For scRNA-seq (R) analysis

Setting up the scRNA-seq data directory

Download data

First download required GSE files (see table at top)

Rename data and put into folders

After downloading all required *barcodes.tsv.gz, *features.tsv.gz, and *matrix.mtx.gz, you will have to rename them and place them into subfolders for the Read10(data.dir=) function. If you place it into a folder called scRNAseq-data it should look like this:

scRNAseq-data
├── 6FDDRR-14dpi-1-iMN
│   ├── barcodes.tsv.gz
│   ├── features.tsv.gz
│   └── matrix.mtx.gz
├── 6FDDRR-14dpi-2-iMN
│   ├── barcodes.tsv.gz
│   ├── features.tsv.gz
│   └── matrix.mtx.gz
├── EmbMN-1
│   ├── barcodes.tsv.gz
│   ├── features.tsv.gz
│   └── matrix.mtx.gz
├── EmbMN-2
│   ├── barcodes.tsv.gz
│   ├── features.tsv.gz
│   └── matrix.mtx.gz
├── LNI-DDRR-4dpi
│   ├── barcodes.tsv.gz
│   ├── features.tsv.gz
│   └── matrix.mtx.gz
├── LNI-DDRR-iMN
│   ├── barcodes.tsv.gz
│   ├── features.tsv.gz
│   └── matrix.mtx.gz
├── LNI-DDRR-nosupplements
│   ├── barcodes.tsv.gz
│   ├── features.tsv.gz
│   └── matrix.mtx.gz
└── MEFs
    ├── barcodes.tsv.gz
    ├── features.tsv.gz
    └── matrix.mtx.gz

Add datadir.txt file to point to data

In the r directory where the .R files are, add a datadir.txt file that points to where the folders with the scRNA-seq data is. For example, if you placed the scRNAseq-data folder on your Desktop, you would put a datadir.txt file in /Users/username/Documents/GitHub/article-engraftable-neurons/r/ that contains the following text

/Users/username/Desktop/scRNAseq-data/

```

This allows the .R scripts to change the working directory to the one containing the data.

### Analysis
Run `2023.10.16_embMNfilter.R` first to isolate the median motor column (MMC) motor neurons. It will produce a .Rds file with just the MMC cells.

Then run `2023.10.17_scRNA-analysis.R`