Cell-Free Gene Expression in Bioprinted Fluidic Networks.

Area of Science:

• Biomaterials Engineering
• Synthetic Biology
• Soft Robotics

Background:

• Developing soft robotic devices with life-like properties necessitates smart materials responsive to environmental cues.
• Cell-free expression systems offer a method for incorporating dynamic molecular control into materials, bypassing complexities of living systems.

Purpose of the Study:

• To integrate cell-free protein synthesis within agarose hydrogels organized by a synthetic vasculature.
• To demonstrate spatial control over protein concentration profiles within these engineered hydrogels.

Main Methods:

• Utilized indirect printing with a fugitive ink (Pluronic F-127) to create fluidic channels in agarose hydrogels.
• Investigated the effect of gel matrix properties (density, expression system dilution) on protein expression from E. coli cell-extract.
• Developed vascularized hydrogels supplied via a single channel to control reactant distribution and protein synthesis.

Main Results:

• Protein expression was found to be dependent on hydrogel density and cell-free system dilution.
• Larger molecules (DNA plasmids) were confined to channels or immobilized, while smaller molecules diffused.
• Demonstrated the ability to generate specific spatial protein concentration profiles (gradients, homogeneous distribution) using different cell-free gene circuits and channel designs.

Conclusions:

• The developed strategy enables the spatial organization and controlled supply of cell-free protein synthesis within hydrogels.
• This approach facilitates the creation of active, responsive soft materials with embedded molecular functions for advanced applications in soft robotics and beyond.