Correlation Between In Vitro and In Vivo Gene-Expression Strengths is Dependent on Bottleneck Process

Constructing gene networks in cells enables the efficient production of valuable substances and the creation of cells performing intended functions. However, the construction of a cellular network of interest, based on a design-build-test-learn cycle, is quite time-consuming due to processes mainly attributed to cell growth. Among the various available methods, cell-free systems have recently been employed for solving network testing problems using cells, because cell-free systems allow quick evaluations of test networks without waiting for cell growth. Although cell-free systems have the potential for use in rapid prototyping platforms, the correlation between the in vitro and in vivo activities for each genetic part (e.g. promoter) remains enigmatic. By quantifying mRNA and its encoded protein in a cell, we have identified appropriate culture conditions where cellular bottlenecks are circumvented and promoter activities are correlated with previous in vitro studies. This work provides a foundation for the development of molecular breadboard research.