Takatoki Yamamoto | PubMed Health

PubMed2006

Controlling the expression ratio of two proteins by inserting a terminator between the two genes.

An approach was developed in controlling the expression ratio of two genes encoded in DNA by inserting a terminator sequence between the two genes. Escherichia coli ribosomal RNA operon T1 terminator, or E. coli tryptophane attenuater was introduced between two green fluorescent protein (GFP) mutants: ultraviolet-optimized mutant (GFPuv: F99S, M153T, V163A) and blue-shifted mutant (BFP: F64L, S65T, T145F). The ratio of the fluorescence intensity of BFP to GFPuv after transcription and translation reflected the termination efficiency of the transcriptional termination activity of the inserted terminator.

Escherichia coli↗

PubMed2002

PDMS-glass hybrid microreactor array with embedded temperature control device. Application to cell-free protein synthesis.

A microreactor array was developed which enables high-throughput cell-free protein synthesis. The microreactor array is composed of a temperature control chip and a reaction chamber chip. The temperature control chip is a glass-made chip on which temperature control devices, heaters and temperature sensors, are fabricated with an ITO (indium tin oxide) resistive material. The reaction chamber chip is fabricated by micromolding of PDMS (polydimethylsiloxane), and is designed to have an array of reaction chambers and flow channels for liquid introduction. The microreactor array is assembled by placing the reaction chamber chip on the temperature control chip. The small thermal mass of the reaction chamber resulted in a short thermal time constant of 170 ms for heating and 3 s for cooling. The performance of the microreactor array was examined through the experiments of cell-free protein synthesis. By measuring the fluorescence emission from the products, it was confirmed that GFP (Green Fluorescent Protein) and BFP (Blue Fluorescent Protein) were successfully synthesized using Escherichia coli extract.

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