Cellulolytic RoboLector - towards an automated high-throughput screening platform for recombinant cellulase...

Area of Science:

• Biotechnology
• Enzymology
• Protein Engineering

Background:

• Cellulases are crucial for cellulose hydrolysis, but their low activity limits biomass conversion efficiency.
• Protein engineering efforts focus on improving cellulase activity through directed evolution and rational design.
• Screening large clone libraries is essential but challenging due to limited automation and reproducibility concerns.

Purpose of the Study:

• To develop and evaluate an extended robotic platform for automated high-throughput screening of cellulase clone libraries.
• To adapt and validate key upstream, downstream, and analytical process steps for automation.
• To ensure reproducible clone ranking for identifying improved cellulase variants.

Main Methods:

• An extended robotic platform was utilized for automated screening, including preculture synchronization and biomass-specific induction.
• Conventional protocols for media preparation, cell lysis, Azo-CMC assay, and PAHBAH assay were adapted for automation.
• The system was evaluated using *E. coli* and *K. lactis* as model organisms.

Main Results:

• Automated protocols for essential process steps were successfully implemented.
• A recombinant *E. coli* celA2 clone library was screened using the automated platform.
• A reliable clone ranking was achieved, demonstrating the system's effectiveness.

Conclusions:

• The RoboLector device is a suitable platform for automated high-throughput screening of cellulase clone libraries.
• On-line biomass growth measurement and controlled liquid handling ensure fair comparison of clone variants.
• The developed system facilitates the identification of improved cellulase enzymes for biomass conversion.