Long-Lasting and Responsive DNA/Enzyme-Based Programs in Serum-Supplemented Extracellular Media

Area of Science:

• Biochemistry
• Synthetic Biology
• Molecular Engineering

Background:

• DNA molecular programs offer versatile biomolecular sensing and actuation for pharmaceuticals.
• Serum nucleases limit DNA program functionality in vitro, causing rapid network deterioration.

Purpose of the Study:

• To develop DNA molecular programs that are functional and stable in serum-rich environments.
• To enhance the biocompatibility and responsiveness of DNA programs for in vitro applications.

Main Methods:

• Implementation of three-letter code DNA networks to suppress parasitic amplification.
• Development of a novel buffer to improve biocompatibility and responsiveness.
• Demonstration of serum-supplemented extracellular DNA programs in the presence of living cells.

Main Results:

• DNA/enzyme programs remained functional in 10% serum for at least 3 days using three-letter code networks.
• The new buffer enhanced biocompatibility and maintained responsiveness to molecular changes.
• Extracellular DNA programs showed rapid responses to molecular inputs (6x faster than cell division) and sustained function over three cell divisions.

Conclusions:

• Developed robust DNA molecular programs functional in serum for over 3 days, overcoming nuclease degradation.
• Demonstrated the potential for in situ biomolecular characterization tools in serum-demanding in vitro models.
• Paved the way for extracellular synthetic biology tools and autonomous in vitro models through chemical reactivity coupling.