Droplet electroporation in microfluidics for efficient cell transformation with or without cell wall removal

Area of Science:

• Biotechnology
• Microfluidics
• Genetic Engineering

Background:

• Cell transformation is crucial for genetic engineering.
• Traditional methods like bulk electroporation have limitations in efficiency and control.
• Microfluidic devices offer potential for improved biological manipulation.

Purpose of the Study:

• To develop and evaluate an efficient cell transformation method using droplet electroporation on a microfluidic chip.
• To compare the efficiency of microfluidic electroporation with bulk electroporation for Chlamydomonas reinhardtii.
• To assess the method's effectiveness for both wild-type and wall-less algal strains.

Main Methods:

• A polydimethylsiloxane-glass microfluidic chip with flow-focusing and serpentine channels was fabricated.
• Droplet generation encapsulated cells and DNA.
• Microelectrode arrays on the chip delivered electroporation pulses.
• Transformation efficiency was measured for wall-less and wild-type Chlamydomonas reinhardtii.

Main Results:

• Microfluidic droplet electroporation achieved over 1000-fold higher efficiency for the wall-less mutant.
• Efficiency was over 100-fold higher for the wild-type Chlamydomonas reinhardtii compared to bulk electroporation.
• The method demonstrated high efficiency even at low DNA/cell ratios, allowing control over transgene copy number.

Conclusions:

• Microfluidic droplet electroporation is a highly efficient and scalable method for algal cell transformation.
• This technique overcomes limitations of bulk electroporation, especially for cell-wall-intact organisms.
• The ability to control transgene copy number enhances the stability of transgene expression.