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Parallel in vivo DNA assembly by recombination: experimental demonstration and theoretical approaches.

Shi Z, Wedd AG, Gras SL - PLoS ONE (2013)

Bottom Line: Despite the availability of computational predictions for well-characterized enzymes, the optimization of most synthetic biology projects requires combinational constructions and tests.A new building-brick-style parallel DNA assembly framework for simple and flexible batch construction is presented here.The assembly of five DNA fragments into a host genome was performed as an experimental demonstration.

View Article: PubMed Central - PubMed

Affiliation: School of Chemistry, University of Melbourne, Parkville, Victoria, Australia. shiz@student.unimelb.edu.au

ABSTRACT
The development of synthetic biology requires rapid batch construction of large gene networks from combinations of smaller units. Despite the availability of computational predictions for well-characterized enzymes, the optimization of most synthetic biology projects requires combinational constructions and tests. A new building-brick-style parallel DNA assembly framework for simple and flexible batch construction is presented here. It is based on robust recombination steps and allows a variety of DNA assembly techniques to be organized for complex constructions (with or without scars). The assembly of five DNA fragments into a host genome was performed as an experimental demonstration.

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Related in: MedlinePlus

The need for topology breakers for circular plasmids.Exchange is shown occurring in the recombination of two linear fragments (shown in red and green) (A). Exchange is also shown for two circular plasmids, which requires an additional recombination (B). Topology breaker sites are shown as triangles.
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pone-0056854-g005: The need for topology breakers for circular plasmids.Exchange is shown occurring in the recombination of two linear fragments (shown in red and green) (A). Exchange is also shown for two circular plasmids, which requires an additional recombination (B). Topology breaker sites are shown as triangles.

Mentions: An assembly unit in a circular carrier (plasmid, phagemid, chromosome) needs an additional recombination site as a topology breaker to ensure that combination between circular conformation plasmids is identical to that between the equivalent linear forms (Figure 5). Two linear DNA molecules exchange parts during recombination, while two circular DNA molecules insert and fuse with each other, i.e., circular DNA molecules will keep fusing if there are no topology breakers.


Parallel in vivo DNA assembly by recombination: experimental demonstration and theoretical approaches.

Shi Z, Wedd AG, Gras SL - PLoS ONE (2013)

The need for topology breakers for circular plasmids.Exchange is shown occurring in the recombination of two linear fragments (shown in red and green) (A). Exchange is also shown for two circular plasmids, which requires an additional recombination (B). Topology breaker sites are shown as triangles.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3585241&req=5

pone-0056854-g005: The need for topology breakers for circular plasmids.Exchange is shown occurring in the recombination of two linear fragments (shown in red and green) (A). Exchange is also shown for two circular plasmids, which requires an additional recombination (B). Topology breaker sites are shown as triangles.
Mentions: An assembly unit in a circular carrier (plasmid, phagemid, chromosome) needs an additional recombination site as a topology breaker to ensure that combination between circular conformation plasmids is identical to that between the equivalent linear forms (Figure 5). Two linear DNA molecules exchange parts during recombination, while two circular DNA molecules insert and fuse with each other, i.e., circular DNA molecules will keep fusing if there are no topology breakers.

Bottom Line: Despite the availability of computational predictions for well-characterized enzymes, the optimization of most synthetic biology projects requires combinational constructions and tests.A new building-brick-style parallel DNA assembly framework for simple and flexible batch construction is presented here.The assembly of five DNA fragments into a host genome was performed as an experimental demonstration.

View Article: PubMed Central - PubMed

Affiliation: School of Chemistry, University of Melbourne, Parkville, Victoria, Australia. shiz@student.unimelb.edu.au

ABSTRACT
The development of synthetic biology requires rapid batch construction of large gene networks from combinations of smaller units. Despite the availability of computational predictions for well-characterized enzymes, the optimization of most synthetic biology projects requires combinational constructions and tests. A new building-brick-style parallel DNA assembly framework for simple and flexible batch construction is presented here. It is based on robust recombination steps and allows a variety of DNA assembly techniques to be organized for complex constructions (with or without scars). The assembly of five DNA fragments into a host genome was performed as an experimental demonstration.

Show MeSH
Related in: MedlinePlus