# Protocol to obtain genetically engineered Acetobacterium woodii and Eubacterium callanderi strains

## TL;DR

This paper provides a detailed protocol for genetically modifying two acetogenic bacteria to convert CO2 and hydrogen into organic acids.

## Contribution

A new protocol for electrocompetent cell preparation and genetic modification of Acetobacterium woodii and Eubacterium callanderi is introduced.

## Key findings

- Electroporation and verification steps for recombinant strains of A. woodii and E. callanderi are described.
- The protocol can be adapted for other anaerobic bacterial strains.
- The method supports the genetic toolbox application for acetogenic bacteria.

## Abstract

We present a protocol to produce electrocompetent Acetobacterium woodii DSM 1030 and Eubacterium callanderi DSM 3468 cells. These acetogens are capable of converting CO2 with H2 and other C1 substrates into organic acids. We describe the electroporation procedure and the verification steps to prove the presence of recombinant strains. This protocol serves as a basis for the application of the genetic toolbox for these bacterial species and can be adapted to several anaerobic strains.

For complete details on the use and execution of this protocol, please refer to Leang et al.1

•Instructions to produce competent anaerobic bacteria cells•Step-by-step protocol for genetic modification of A. woodii and E. callanderi•Overview of common difficulties in producing recombinant strains

Instructions to produce competent anaerobic bacteria cells

Step-by-step protocol for genetic modification of A. woodii and E. callanderi

Overview of common difficulties in producing recombinant strains

Publisher’s note: Undertaking any experimental protocol requires adherence to local institutional guidelines for laboratory safety and ethics.

We present a protocol to produce electrocompetent Acetobacterium woodii DSM 1030 and Eubacterium callanderi DSM 3468 cells. These acetogens are capable of converting CO2 with H2 and other C1 substrates into organic acids. We describe the electroporation procedure and the verification steps to prove the presence of recombinant strains. This protocol serves as a basis for the application of the genetic toolbox for these bacterial species and can be adapted to several anaerobic strains.