Team:ZJU-China/Design

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Circuit design

Lack a brief intro of this page.

Support device(Click different parts on figure below to learn more )

1.BBa_K1433001 attB-BBa_J23110-attP

BBa_J23110 is a middle strength promoter from the constitutive promoter family, which are flanked by attB and attP sites, constructing the core parts of bistable switch. The direction of the promoter, which is controlled by Int and Xis, decides the state of support device is controlled (See bistable switch page for detailed information.)int-xis-bplr.gif

Socket

Socket is a circuit constructed on host cells, which contains homology regions and int gene. All construction happens in this circuit.(Click different parts on figure below to learn more)

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1. Promoter BBa_J23110

The promoter we use here has two functions:

  1. To promote transcription of Int after bi-terminator unit is replaced by the inserted circuit.
  2. To promote the inserted circuit CDS if needed.

BBa_J23110 is a middle strength promoter from a constitutive promoter family. It can be replaced if users do not need it, but users should notice that there is a promoter behind bi-terminator unit which allows Int expression. (See solutions below)

Socket.coli

Socket.coli is the final product of our project. It’s the host of GeneSocket.

In Socket.coli, the Socket circuit is inserted into bacterial chromosome in advance by using existed chromosome recombination system. The bi-terminator unit is the first site waiting for parts to be inserted. Support device is also transformed into the cell. The original state of bistable switch is WORK state, waiting for the first part to be inserted.

With this strain, GeneSocket can work as this page shows. The cell has at least 3 functions:

  1. Store GeneSocket system.
  2. Be the object of GeneSocket recombination.
  3. Easily amplify GeneSocket and spread it as a product!

Easily amplify GeneSocket and spread it as a product!

The main function of standard parts is adding the next bi-terminator unit into SOCKET together with current insertion parts, which is a crucial design for continuously circuit construction. Besides, since the circuit situation can be quite different, we provide two parts to help you deal with most cases.

1. Homology region of inserted part.

The homology regions flanking beside inserted parts are added by PCR. The Latter parts must have SpeI restriction site to link with our standard parts.

References


[1]Bonnet, J., Subsoontorn, P. & Endy, D. Rewritable digital data storage in live cells via engineered control of recombination directionality. Proceedings of the National Academy of Sciences of the United States of America 109, 8884-8889, doi:10.1073/pnas.1202344109 (2012).

[2]Wargacki, A. J. et al. An engineered microbial platform for direct biofuel production from brown macroalgae. Science 335, 308-313, doi:10.1126/science.1214547 (2012).