Team:Emory/Description

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DESCRIPTION

Synthetic Biology is largely restricted to well-funded laboratories at major research universities in high income countries. One significant barrier to entry is the capital cost of instruments. The cloning and assembly of BioBricks, for example, includes the transformation of Escherichia coli, which requires the purchase of a refrigerated centrifuge and an ultra-cold freezer. Here we assemble BioBrick-compatible shuttlevectors for Acinetobacter baylyi ADP1, a naturally competent relative of E. coli that grows as rapidly under identical conditions. We will show that A. baylyi can be transformed with recombinant DNA simply by adding ligation reactions to mid-log cultures; transformants are selected as usual by spreading them onto LB agar plates supplemented with the appropriate antibiotics (kanamycin, spectinomycin, tetracycline, cefotaxime or amikacin). These experiments will show how BioBricks can be constructed and assembled in modestly funded laboratories in community colleges, high schools and even private homes. The resulting plasmid constructs retain their pSB1C3 backbones and will thus remain compatible with the BioBrick standard and capable of replication in the widely used E. coli chassis.

What is the context of this research?

Acinetobacter baylyi ADP1 grows under the same conditions as E. coli at comparable rates, but requires no encouragement to import DNA from its environment. The only catch is that plasmids that replicate in E. coli don't also replicate in A. baylyi. We are creating plasmids that replicate in both species to harness A. baylyi for genetic engineering.

What is the significance of this project?

The experiments proposed here will show the that Acinetobacter baylyi ADP1 can be transformed with foreign DNA cloned into our shuttle vectors, thus obviating the need for expensive equipment to perform a simple transformation. We plan to make our genetic tools freely accessible to modestly funded labs in community colleges, high schools and private homes. They could also be used to streamline workflows at well-funded labs.

What are the goals of the project?

We are constructing shuttle vectors for Acinetobacter baylyi ADP1, a naturally competent relative of E. coli that grows rapidly under identical culture conditions. We will add ligation reactions containing small amounts of cut and pasted DNA to mid-log cultures of A. baylyi. The resulting bacterial culture will be spread onto Lysogeny Broth agar plates supplemented with antibiotics to select against untransformed cells. If the transformation is successful, we will see the presence of antibiotic-resistant bacterial colonies. These colonies will prove that we can transform bacteria without effort or expensive equipment.