Cell-free transcription-translation (TX-TL) systems have been developed in the 60s to study the process of protein synthesis and gene regulation in bacteria. After the invention of highly efficient cell-free systems in the 90s, in vitro protein synthesis became widely used for applications in biotechnology, industry and proteomics, as an alternative to the recombinant protein technology. In the past decade, it has been suggested that cell-free TX-TL systems could also become powerful platform for synthetic biology purposes.I will present a cell-free TX-TL toolbox specifically developed to program complex biochemical processes in vitro. This system uses the endogenous E. coli TX-TL machineries to construct and to study elementary gene circuits in test tubes. Recently we showed that the expression of large DNA programs could be also carried out with this system. The bacteriophage T7 can be entirely synthesized in vitro from its genomic DNA. In addition, we show that the replication of the DNA instructions occurs concurrently with the phage expression and self-assembly. This work is not limited to T7, as the phage phiX174 is also entirely synthesized with the same system and in the same conditions. I will discuss the advantages, limitations and perspectives of this approach.

Molecular programming with a cell-free transcription-translation system: Elementary gene circuits and phage synthesis in a test tube

244th National Meeting (2012)

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