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From 2014.igem.org

Overview

In order to achieve the goal of integrating the directed evolution processes into a single automatic and controllable system, we come up with the following design.

1.The initiate state

M13 bacteriophage with gⅧ deficient carrying Prey sequence yet to be evolved. An E. coli with pBT-Bait and pRPT-gⅧ. At the temperature of 0℃, M13 phage only attach but not infect into the cell.

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2.Infection

When the temperature rise to 30℃, the M13 vector infects into the E. coli.

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3.Mutagenesis

When the M13 vector undergo replication, a library of different Prey is generated.

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4.Selection

Various Prey undergo selection processes with the help of Bacteria Two-Hybrid system.Only the Prey that can have interaction with Bait can activate transcriping of gⅧ.

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5.Temperature regulated amplification

The temperature rise to 37℃, so that mRNA of gⅧ can be translated into p8.

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6. Rescue and Evolved

The p8 compensate M13 deficient, so that progeny phage can be generated.

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