Device

Part:BBa_K1412000

Designed by: Chun Tang, Jielin Chen, Yue Jiang   Group: iGEM14_XMU-China   (2014-08-16)

CheZ generator under Lac promoter


This part consists of Lac promoter and CheZ generator. It could endow chemotaxis ability to CL-1.

Usage and Biology

CheZ can dephosphorylate CheY-P to CheY. CheY could make the flagellar motor rotate counterclockwise so that the cell could swim. Without CheZ, CheY-P is phosphorylated, which could bind to the flagellar motor protein FliM, causing the cell to tumble. The expression of CheZ allows E.coli to navigate gradients of natural chemoattractants.

At the same time, we know that Lac promoter is an inverting regulator sensitive to LacI and CAP, while LacI can be inhibited by IPTG. With LacI protein, CAP protein and IPTG, we can adjust the expression of CheZ via Lac promoter to control the movement of E.coli. The part BBa_K1412001 has already been used to govern E.coli chemotaxis successfully.


Sequence and Features


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    COMPATIBLE WITH RFC[12]
  • 21
    COMPATIBLE WITH RFC[21]
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    COMPATIBLE WITH RFC[25]
  • 1000
    COMPATIBLE WITH RFC[1000]

Experimental Data

CL-1 and BBa K1412000.png

Figue 1. The comparison of chemotaxis ability between device BBa_J04450 and BBa_K1412000

CheZ and LacI are both knocked out of the genome of CL-1, thus CL-1 does not have the ability of chemotaxis and will not inhibit the expression of Lac promoter. Therefore CL-1 could express CheZ in BBa_K1412000 to regain chemotaxis ability. While compare with BBa_J04450, which has not the ability of chemotaxis. We can get the result shown on the figure, the diameter of chemotaxis: d2 > d1. It mean that BBa_K1412000 have the chemotaxis ability.

Protocol

We used M63 semi-solid plate to observe the behavior of the two E.coli(CL-1) strain. We spotted 3μL of the cells on the same plate, then cultured the plate at 37℃ incubator. By comparing the chemotaxis diameter of the bacteria on the plate, we conclude that BBa_K1412000 could endow chemotaxis ability to CL-1.

References

[1] [http://pubs.acs.org/doi/abs/10.1021/ja0692480 Topp S, Gallivan J P. Guiding bacteria with small molecules and RNA[J]. Journal of the American Chemical Society, 2007, 129(21): 6807-6811.].

More information, click here: [http://2014.igem.org/Team:XMU-China# XMU-China]

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