Part:BBa_I716462:Experience

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Applications of BBa_I716462

Datasheet for Part BBa_I716462 in E. coli strain DH5alpha.

Characterization by Lethbridge iGEM Team 2011

Main Introduction

Part I716462 was previously characterized by the UC Berkeley iGEM team in 2007. The purpose of this part was to prevent unwanted cell proliferation by engineering a genetic self-destruct mechanism into bacteria that upon induction would express a genetic material-degrading toxin, the endonuclease BamHI that would kill cells yet preserve their physical integrity. BamHI has a restriction cut site of G'GATCC, generating sticky ends (1). The restriction endonuclease was placed under the control of the arabinose- inducible promoter, pBAD. In 2007, the UC Berkeley iGEM team was able to show that cells lost their ability to reproduce but did not lyse upon induction with arabinose (2). The University of Lethbridge 2011 iGEM team wished to further characterize this part to degrade the genomic DNA of E. coli in order to have a functional chassis for our tailings pond clean-up kit without the risk of DNA contamination. In this way, tight control and regulation over our engineered bacterial cells is possible.

Characterization

Cell Survival

Materials and Methods

The BamHI construct was overexpressed in E. coli DH5α cells. Cultures were induced with arabinose at an OD₆₀₀ of 0.6 and grown overnight. Samples of the induced culture and an uninduced culture were plated on LB agar plates to monitor growth of colony forming units.

References

(1) Viadiu H, Aggarwal AK (2000). Structure of BamHI bound to nonspecific DNA: a model for DNA sliding. Mol. Cell 5 (5): 889-895.
(2) iGEM 2007. (2011, May 10). BerkiGEM2007Present5. Retrieved from http://parts.mit.edu/igem07/index.php/BerkiGEM2007Present5.

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