2019

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Team of Alive

Abstract

There is an increasing demand in biomedical research for techniques to monitor the dynamics of multiple genes over several time points. However, current methods such as gene reporters are limited to a few genes of interest or require sample destruction in the case of transcriptomic analysis. We thus engineered ALiVE as a diagnostics platform for the Analysis of Living cells via Vesicular Export. In particular, we hijacked the mechanisms of exosome secretion and viral budding to export specific transcripts from living cells repeatedly over time. Based on versatile BioBricks, we modified membrane proteins with affinity tags which are fused to bio-orthogonal RNA-adapters to enable convenient purification of the vesicles and thus the exported RNA. We also introduced sensitive luciferase reporters to quantify vesicle secretion efficiency and collateral transfection. ALiVE is a generalizable technology for minimally invasive diagnostics of gene expression dynamics in cellular model systems and holds great promise for monitoring cellular therapies in regenerative medicine.

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