Automated Illumina NGS Library Prep with PyLabRobot

This repository contains scripts for running automated Illumina NGS library preparation on a Hamilton STARlet liquid handler using PyLabRobot. This method originally was designed for the Illumina NGS Library Prep workflow at Pioneer Labs.

The full workflow includes:

• Sample consolidation — consolidating samples across 96-well plates
• Adapter PCR — running adapter PCR with the same primers across all samples
• Index PCR — running index PCR with different primers per sample

Each step can also be run independently.

Method Overview

Recommended workflow

1. Read docs/LPM_USERGUIDE.md to learn more about the workflow
2. Follow installation instructions below to set up your environment and install PyLabRobot
3. Execute method

Prerequisites

• Hardware: Hamilton STARlet liquid handler (required for real runs; simulation mode is available without hardware — see methods/startup.py)
• Python: 3.12
• PyLabRobot: Version 0.2 as of March, 2026 (see PyLabRobot Resources below)
• Package manager: conda (recommended) or pip/venv
• Slack workspace: optional, for run notifications

Installation

1. Clone the repository

git clone https://github.com/your-org/ngs-library-prep.git
cd ngs-library-prep

2. Set up the environment

Choose either conda (recommended) or venv.

Option A — conda (environment.yml)

conda env create -f environment.yml
conda activate ngs_library_prep

Option B — venv (requirements.txt)

python3 -m venv .venv
source .venv/bin/activate        # macOS/Linux
# .venv\Scripts\activate         # Windows
pip install -r requirements.txt

3. Clone PyLabRobot (following User Guide on PyLabRobot.org) inside the ngs_library_prep repo

git clone https://github.com/pylabrobot/pylabrobot.git
cd pylabrobot
pip install -e '.[dev]'

Scripts

Notebook	When to use
scripts/library_prep.ipynb	Full workflow: lysis resuspension → sample consolidation → adapter PCR setup
scripts/adapter_to_index_PCR.ipynb	Mid-workflow entry: adapter PCR → optional Exo1 dispense → index PCR

Repository Structure

ngs_library_prep/
├── docs/                        # Documentation and user guides
│   ├── LPM_USERGUIDE.md         # Step-by-step user guide for the full workflow
│   └── imgs/                    # Images referenced by the user guide
├── methods/                     # Core Python functions for each workflow step
│   ├── library_prep.py
│   └── startup.py
├── resources/                   # Custom labware definitions
│   ├── __init__.py
│   └── custom_labware.py
├── scripts/                     # Jupyter notebooks for executing workflows on the robot
│   ├── library_prep.ipynb
│   └── adapter_to_index_PCR.ipynb
├── utils/                       # Shared utility helpers
│   ├── labware_utils.py
│   └── robot_utils.py
├── worklists/                   # Example input worklists
│   └── example_worklist.csv
├── environment.yml              # Conda environment spec
├── requirements.txt             # pip/venv dependency list
└── example.config.json          # Example configuration file for Slack API integration

Resources for Pioneer Labs

• Website
• Substack
• Issues? Add via Github Issues

Resources for PyLabRobot

Documentation

docs.pylabrobot.org

Support

• discuss.pylabrobot.org for questions and discussions.
• GitHub Issues for bug reports and feature requests.