Composite

Part:BBa_K1212013

Designed by: Aura Ferreiro   Group: iGEM13_UC_Davis   (2013-09-16)

pTet + TBS1 + RBS + GFP

See our [http://2013.igem.org/Team:UC_Davis/Data wiki's data page] for more information.
pTet promoter upstream of TALe binding site 1 (corresponding to TAL 1 [http://partsregistry.org/wiki/index.php?title=Part:BBa_K1212004 (BBa_K1212004)] , with B0034 and Golden Gate compatible GFP.
The graph below shows GFP fluorescence normalized by OD 600 readings over the course of a bit less than 15 hours with pTet induced by 100 ng/mL aTc and not induced. There is little to no fluorescence when no inducer is added and a 74 fold increase in fluorescence is seen when induced with 100 ng/mL aTc. This experiment was done in E.coli K-12 strain MG1655Z1, which has been engineered to overexpress TetR.

TBS1_GFP_graph_UCDavis.jpg


The data below shows the behavior of this part in the context of a cotransformation between pBAD+RS2+TAL 1 in pSB3K3 (low copy number) and pTet+TBS 1+GFP in pSB1C3 (high copy number). Also see [http://partsregistry.org/wiki/index.php?title=Part:BBa_K1212011 BBa_K1212011] and [http://partsregistry.org/wiki/index.php?title=Part:BBa_K1212014 BBa_K1212014] for further characterization of our RiboTALes.

UCD_2013_RiboTALE_Tunability.PNG



The image above displays the peak fluorescence of two RiboTALe constructs, one expressing TALe 1 and the other expressing TALe 8, under different induction conditions for arabinose and theophylline. Both RiboTALes exhibit the expected behavior pattern given the induction conditions, but at consistently different levels of fluorescence. We have attributed this to the difference in binding affinities of the two TAL repressors to their respective binding sites.This variable, if well characterized for different TAL repressors, will provide a powerful means to control the tunability of these devices.


TBS sequence obtained from
J. F. Meckler, M. S. Bhakta, M. S. Kim, R. Ovadia, C. H. Habrian, A. Zykovich, et al., "Quantitative analysis of TALE-DNA interactions suggests polarity effects," Nucleic Acids Res, vol. 41, pp. 4118-28, Apr 2013.


Sequence and Features


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    INCOMPATIBLE WITH RFC[12]
    Illegal NheI site found at 125
  • 21
    INCOMPATIBLE WITH RFC[21]
    Illegal BamHI site found at 65
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    COMPATIBLE WITH RFC[25]
  • 1000
    COMPATIBLE WITH RFC[1000]


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Categories
Parameters
None