Seeing the Invisible, and Considering Context: Exploiting Crosstalk to Measure Genetic Construct Activity

â??Seeing the Invisible, and Considering Context: Exploiting Crosstalk to Measure Genetic
Construct Activity�
Ariel Hecht, Matthew S. Munson, Drew Endy, and Marc Salit
The ability to quantify the activity of genetic parts that do not produce spectroscopically quantifiable products would support the reusability of parts in a forward engineering context. A potential solution, described here, is a competitive assay performed in an in vitro transcription- translation (IVTT) system. In an in vitro environment, expression of orthogonal genes are coupled by competition for limited resources, which can be exploited for measuring the activity of â??invisibleâ? genetic constructs. Several new results from previously presented work are highlighted here:
1. The activity of the non-fluorescent construct of interest can be measured by exploiting resource-based coupling in a competitive assay.
2. A linear relationship between the activities of two constructs is predicted only under specific conditions.
3. The relative activities of constructs are context-dependent: assay measurements cannot yet be ported between expression systems.
The above results suggest that, while the competitive assay is feasible, appropriate conditions that define a context must be identified. Future work will focus on identifying assay conditions using a validated model-based framework, potentially leading to a procedure for characterizing the capacity of both in vitro and in vivo expression contexts.