Controlled exchange of protein and nucleic acid signals from and between synthetic minimal cells.

Synthetic minimal cells are a class of bioreactors that have some, but not all, functions of live cells. Here, we report a critical step toward the development of a bottom-up minimal cell: cellular export of functional protein and RNA products. We used cell-penetrating peptide tags to translocate payloads across a synthetic cell vesicle membrane.

Programmable Fusion and Differentiation of Synthetic Minimal Cells.

Synthetic cells can mimic the intricate complexities of live cells, while mitigating the level of noise that is present natural systems; however, many crucial processes still need to be demonstrated in synthetic cells to use them to comprehensively study and engineer biology. Here we demonstrate key functionalities of synthetic cells previously available only to natural life: differentiation and mating. This work presents a toolset for engineering combinatorial genetic circuits in synthetic cells.

Liposome Preparation by 3D-Printed Microcapillary-Based Apparatus.

Liposomal encapsulation serves as the basis for the engineering of biomimetic and novel synthetic cells. Liposomes are normally formed using such methods as thin film rehydration (TFH), density-mediated reverse emulsion encapsulation (REE), or one of many microfluidics-based approaches-with the latter of these two methods being used mainly for the encapsulation of various lumen constituents such as cell-free protein expression reactions. Here, we describe the simultaneous formation and encapsulation of liposomes and various cell-mimetic lumen chemistries, respectively, using a 3D-printable microcapillary-based microfluidics device based off of the droplet-shooting and size-filtration (DSSF) liposome preparation method.

Methods for thermal denaturation studies of nucleic acids in complex with fluorogenic dyes.

Thermal denaturation is a common technique in the biophysical study of nucleic acids. These experiments are typically performed by monitoring the increase in absorbance (hyperchromism) of a sample at 260nm with temperature (Mergny & Lacroix, 2003; Puglisi & Tinoco, 1989). This wavelength is chosen as nucleic acids of mixed sequence typically exhibit their maximum absorbance here.