Rapid Development of Cell State Identification Circuits with Poly-Transfection.

Researchers from the Massachusetts Institute of Technology (MIT) and the University of British Columbia have developed a groundbreaking technique called poly-transfection, which is an extension of traditional mammalian transfection. This new method involves subjecting each cell within a transfected population to a unique experiment, allowing for the analysis of numerous DNA copy numbers in a single reaction. Poly-transfection has been successful in optimizing the ratios of three-component circuits within a single cell well, with the potential for application to even larger circuits.

The outcomes of this innovative method have practical implications in determining optimal DNA-to-co-transfect ratios for transient circuits and selecting expression levels for circuit components when generating stable cell lines. The researchers have demonstrated the application of poly-transfection in optimizing a three-component circuit, starting with experimental design principles and building upon traditional co-transfection methods. Flow cytometry is conducted several days after poly-transfection, and the resulting data analysis examines specific segments of single-cell flow cytometry data corresponding to cells characterized by distinct component ratios.

Poly-transfection has proven useful in laboratory settings for optimizing various genetic circuit elements, including cell classifiers, feedback and feedforward controllers, and bistable motifs. This method has the potential to significantly accelerate the design cycles associated with complex genetic circuits in mammalian cells, leading to advancements in research and applications in the field. The researchers have published their findings in the Journal of Visualized Experiments, which provides a detailed protocol for poly-transfection.

doi: 10.3791/64793.