Title: Data for: ALK R1275Q mutation drives expansion of SCP-like cells during sympathoadrenal commitment and primes neuroblastoma initiation
DOI: https://doi.org/10.48723/9bm1-gr40

# RNA-seq Raw Data

This directory contains raw sequencing data from both bulk RNA-seq and single-cell RNA-seq experiments.

## Directory Structure

```
.
├── BulkRNA.d/        # Bulk RNA-seq data (FASTQ files)
├── scRNA.d/          # Single-cell RNA-seq data (Cell Ranger outputs)
└── README.md         # README file, this file
```

---

## Bulk RNA-seq Data (`./BulkRNA.d/`)

### Overview

This directory contains raw sequencing data (FASTQ format) from bulk RNA-seq experiments.


### Sample Information

#### Sample Types
| Type | Description |
|------|-------------|
| Ctrl | Healthy donner samples |
| NB   | Neuroblastoma patient donner samples |

#### Sample IDs
- Ctrl7, Ctrl10, Ctrl14
- NB1, NB2, NB4

#### Treatment Conditions
| Condition | Description |
|-----------|-------------|
| D6, D8, D10, D12 | Different time points (days) in sympathoadrenal mature stage |
| NCC | Neural Crest Cells stage |
| SAP | Sympathoadrenal progenitor stage |

### File Naming Convention

Format: `{SampleID}_{Condition}_{ReadNumber}.fq.gz`

- `1.fq.gz` - Forward reads (Read 1)
- `2.fq.gz` - Reverse reads (Read 2)

### Example
```
Ctrl10_D10_1.fq.gz  - Ctrl10 sample, D10 condition, forward reads
Ctrl10_D10_2.fq.gz  - Ctrl10 sample, D10 condition, reverse reads
```

### Data Summary

| Sample | Conditions | Total Files |
|--------|-----------|-------------|
| Ctrl7  | D6, D8, D10, D12, NCC, SAP | 12 |
| Ctrl10 | D6, D8, D10, D12, NCC, SAP | 12 |
| Ctrl14 | D6, D8, D10, D12, NCC, SAP | 12 |
| NB1    | D6, D8, D10, D12, NCC, SAP | 12 |
| NB2    | D6, D8, D10, D12, NCC, SAP | 12 |
| NB4    | D6, D8, D10, D12, NCC, SAP | 12 |
| **Total** | - | **72 files** |

### File Format

- **Format**: FASTQ compressed with gzip
- **Extension**: .fq.gz
- **Sequencing**: Paired-end (2 reads per sample)
- **Date**: March 14, 2023

---

## Single-cell RNA-seq Data (`./scRNA.d/`)

### Overview

This directory contains raw sequencing data and Cell Ranger processing outputs for single-cell RNA-seq experiments with hashtag oligo (HTO) multiplexing.

### Directory Structure

```
scRNA.d/
├── 10X_23_051/    - Sample 1 data and Cell Ranger output
├── 10X_23_052/    - Sample 2 data and Cell Ranger output
├── 10X_23_053/    - Sample 3 data and Cell Ranger output
├── 10X_23_054/    - Sample 4 data and Cell Ranger output
├── V350202566/    - Original raw sequencing data by lane
└── library_indices.tsv
```

### Samples Overview

| Sample ID | Barcode R1 | Barcode R2 | Purpose |
|-----------|------------|------------|---------|
| 10X_23_051 | GTGCTTATCA | CATCCAGCCG | Gene Expression + HTO |
| 10X_23_052 | ACTCGATGCA | CTCGCCCTAT | Gene Expression + HTO |
| 10X_23_053 | AGAAACACAA | GCTTCTTGCT | Gene Expression + HTO |
| 10X_23_054 | GAAGTTAGGG | TCACGCGTTA | Gene Expression + HTO |

### Sample Directory Contents

Each `10X_23_0XX/` directory contains:

- `*_GE_S1_L001_R1_001.fastq.gz` - Gene Expression read 1 (cell barcode + UMI)
- `*_GE_S1_L001_R2_001.fastq.gz` - Gene Expression read 2 (cDNA)
- `*_HTO_S1_L001_R1_001.fastq.gz` - HTO read 1 (cell barcode + UMI)
- `*_HTO_S1_L001_R2_001.fastq.gz` - HTO read 2 (hashtag)
- `config.csv` - Cell Ranger multi configuration file
- `10X_23_0XX/` - Cell Ranger multi output directory
  - `outs/` - Main output directory
    - `per_sample_outs/` - Sample-specific outputs
      - `count/` - Count matrix (features.tsv.gz, barcodes.tsv.gz, matrix.mtx.gz)
      - `metrics_summary.csv` - QC metrics
      - `web_summary.html` - Interactive QC report

### Original Raw Data (V350202566/)

The `V350202566/` directory contains original Illumina sequencing data before demultiplexing:

- `L01/` - Lane 1 raw data
- `L02/` - Lane 2 raw data
- `L03/` - Lane 3 raw data
- `L04/` - Lane 4 raw data

### Data Processing Pipeline

1. **Demultiplexing**: Original fastq files (V350202566) were demultiplexed using SplitDualBarcodes.pl based on dual barcode indices in mingzhi_library_indices.tsv
2. **Cell Ranger multi**: Each demultiplexed sample was processed using Cell Ranger multi with configuration in config.csv
3. **Downstream Analysis**: Processed data for further Seurat-based analysis

### Files

- `library_indices.tsv` - Dual barcode indices for each sample (GE and HTO)

### Data Format

- Fastq files: gzip-compressed FASTQ format
- Count matrices: 10x Genomics mtx format (features.tsv.gz, barcodes.tsv.gz, matrix.mtx.gz)
- Metrics: CSV and HTML format for QC visualization

---

## Notes

- All files are gzip-compressed
- Bulk RNA-seq data date: March 14, 2023
- Single-cell RNA-seq data date: October 31, 2024


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