Understanding the genetic basis of adaptive phenotypes is a central goal in modern ecology and evolutionary biology. The CRISPR-Cas9 genome editing system is a flexible, accessible, and powerful tool that has become standard in functional genomics. Combined with ‘omics’ technologies, CRISPR enables researchers to more readily investigate the links between genotype and phenotype in non- model (or so-called ‘specialist’) species, helping to reveal new insights across the tree of life. In this course, participants will learn how to design and implement CRISPR-Cas9 gene knockout (KO) experiments. Through a mix of lectures and project-based hands-on activities, the course will provide both theoretical foundations and practical guidance for successful Cas9-mediated KOs in an evolutionary context.

This course is designed for undergraduate and graduate students, researchers, and technicians working in biology and ecology, who wish to implement CRISPR-Cas9 tools in their projects. Drawing on the instructor's background in functional genomics of non-model insects, we will focus on the application of CRISPR in evolutionary and ecological studies using examples from insect molecular biology. Nonetheless, the principles of experimental design and methods covered in this course are broadly transferable to other biological systems, with appropriate species-specific adjustments.

A basic background in genetics and molecular biology is required, and participants should have a solid understanding of the central role of nucleic acids (DNA and RNA) in evolution and adaptive phenotypes. We also assume basic familiarity with the UNIX environment. If you are new to UNIX, please complete this tutorial before the course begins. Complementary terminal commands will be provided during the course as needed.

This course will combine theoretical lessons with hands-on demonstration sessions. To accommodate diverse research interests, the course will emphasize experimental design, method selection, and data analysis. 

A dedicated Slack workspace will be set up to support class interactions, and all materials and code examples will be available on a dedicated GitHub repository.

Special time will be reserved for personal project discussions, giving participants the opportunity to share and improve their planned or ongoing projects with instructor guidance.

By the end of the course, participants will be able to:

• Understand the fundamentals of the CRISPR-Cas9 genome editing tool.

• Design a reliable CRISPR-based gene knockout (KO) experiment.

• Design and evaluate species-specific single guide RNAs (sgRNAs).

• Efficiently synthesize sgRNAs and assemble optimized Cas9/sgRNA mixes.

• Screen for CRISPR-derived mutants using a number of genotyping methods.

• Analyze Cas9-induced genomic variants by Illumina deep sequencing.

• Critically interpret the results from a CRISPR-based KO experiment

Daily schedule: from 3 – 8 pm Berlin time.

DAY 1

• Welcome and course overview

• The history of CRISPR-Cas9 technology

• Case Study 01: A showcase of CRISPR-Cas9 for evolution and ecology studies

• Presentation to the course hands-on project

• (hands-on) On reference sequences and gene annotations

• Open Q&A

DAY 2

• Experimental considerations of Cas9-based KO experiments

• Key considerations for designing optimal sgRNAs

• (hands-on) Designing custom sgRNAs pt. 1: Web-based

• (hands-on) Analyzing sgRNA oligos

• Open Q&A

DAY 3

• (hands-on) Designing custom sgRNAs pt. 2: UNIX-based

• Synthesis of sgRNAs via in vitro transcription

• Methods for evaluating sgRNA efficiency

• Case Study 02: Optimizing the delivery of CRISPR components via microinjections

• Open Q&A

DAY 4

• Screening for Cas9-induced mutations

• Genotyping Cas9-modified individuals

• (hands-on) Analyzing Illumina data from KO mutants

• Conclusion of course hands-on project

• Open Q&A

Instructor

Cancellation Policy:

> 30  days before the start date = 30% cancellation fee

< 30 days before the start date= No Refund.

Physalia-courses cannot be held responsible for any travel fees, accommodation or other expenses incurred to you as a result of the cancellation.