# Efficient CRISPR-Cas genome editing in brown algae

## TL;DR

Researchers developed a CRISPR-based genome editing method for brown algae that is efficient, transgene-free, and works across multiple species.

## Contribution

A high-efficiency, transgene-free CRISPR platform for genome editing in brown algae, applicable across multiple species.

## Key findings

- PEG-mediated RNP delivery enables reproducible editing in Ectocarpus without cloning or specialized equipment.
- Precise editing of the IMM locus recreated the imm mutant phenotype, demonstrating platform effectiveness.
- The method is transferable to other brown algal species, including kelps, enabling functional genomics studies.

## Abstract

Brown algae represent the third most complex lineage to have independently evolved multicellularity, distinct from plants and animals. Yet, functional studies of their development and evolution have been limited by the absence of efficient genome editing tools. Here, we present a robust, high-efficiency, and transgene-free CRISPR-based genome editing platform applicable across four ecologically and biotechnologically important brown algal species. Using Ectocarpus as a model, we optimized a polyethylene glycol (PEG)-mediated ribonucleoprotein (RNP) delivery system that achieves reproducible editing across multiple loci without cloning or specialized equipment. As proof of concept, we recreated the hallmark imm mutant phenotype by precisely editing the IMMEDIATE UPRIGHT (IMM) locus. APT/2-fluoroadenine (2-FA) selection further enhanced specificity with minimal false positives. The method was easily transferable to other species, including kelps. This platform now enables functional genomics in brown algae, providing powerful tools for investigating development, life cycle regulation, and the independent evolution of complex multicellularity.

•We develop high-efficiency, transgene-free CRISPR genome editing for brown algae•We optimize PEG-RNP delivery to achieve reproducible editing across multiple loci•We demonstrate precise editing of the imm locus in Ectocarpus•We provide a platform to enable functional genomics in brown algae

We develop high-efficiency, transgene-free CRISPR genome editing for brown algae

We optimize PEG-RNP delivery to achieve reproducible editing across multiple loci

We demonstrate precise editing of the imm locus in Ectocarpus

We provide a platform to enable functional genomics in brown algae

Although most of the biodiversity on Earth lives in oceans, a significant proportion of its organisms remain largely uncharacterized. Brown algae represent one such understudied group of marine photosynthetic eukaryotes. Despite their importance as emerging models for developmental evolution and blue biotechnology, functional genomics in brown algae has remained largely inaccessible due to a lack of efficient and scalable genome editing tools. Our aim is to democratize genome editing in brown algae by developing a high-efficiency, transgene-free protocol that works across multiple species, without the need for specialized equipment.

Brown algae, a key yet understudied lineage of multicellular eukaryotes, have long lacked tools for functional genomics. Martinho et al. introduce a high-efficiency, transgene-free CRISPR platform enabling precise genome editing across multiple brown algal species, unlocking new possibilities for studying their development, evolution, and biotechnological potential.