Automating transformation in non-model bacteria in 96-well format

Classical model organisms such as E. coli and B. subtilis benefit from decades of tool development, standardized protocols, and reliable genetic engineering methods. Non model bacteria, on the other hand, often lack these resources - even though many display phenotypes of high industrial and environmental value, such as plastic degradation, plant growth promotion and biopolymer production.
 

In a recent webinar, Andrea M. Garza Elizondo, PhD, from Oak Ridge National Laboratory (ORNL) described how automation and electroporation optimization can dramatically accelerate genetic transformation workflows for these hard-to-engineer microbes*.

Bacterial Genetic Engineering

Successfully engineering bacteria requires overcoming three major hurdles:

1. Introduction – Getting DNA, and potentially other cargos, efficiently into the cell
2. Maintenance – Ensuring the DNA is retained
3. Function – Achieving reliable gene expression

To address this challenge, the ORNL team set a clear goal: develop an automated script that identifies the most efficient electroporation pulse for transforming plasmid DNA into a given bacterium.

Leveraging existing automation infrastructure - including the Lonza 4D Nucleofector 96-well Unit, liquid handling robots, plate readers, incubators, and colony pickers - the team designed a workflow that:

• Tests 17 bacterial pulse codes simultaneously
• Includes replicates and no-pulse controls
• Automates serial dilution and spot plating
• Compares survival and transformation efficiency using ± antibiotic selection

In this workflow, the team examined key sources of error, such as: Pipetting accuracy during serial dilutions (especially, when reusing pipette tips), dispensing height during spot plating and effects of agar unevenness, liquid viscosity, and dispensing speed.

Rather than setbacks, these findings highlighted the strength of automation: errors became visible, reproducible, and therefore correctable.

By automating the entire workflow, this approach enabled a fast transition from non model bacteria to engineered strains, minimizing manual effort and variability. This allowed researchers to focus on bacterial phenotypes rather than strain handling, with the ultimate goal of a fully integrated, high throughput platform covering:

• Inoculation and growth in 96 well plates
• Timed addition of cell wall weakening agents
• Harvesting, washing, and buffer exchange
• DNA addition, electroporation, and recovery
• Automated dilution, plating, incubation, and analysis

Discover how automated electroporation reshapes bacterial transformation on our dedicated application page including helpful resources, and watch the on-demand webinar. If you have any further questions, our Scientific Support Team at Lonza will be happy to assist you.

Written by 

Isa
Senior Scientific Support Specialist