Device

Part:BBa_M36532:Design

Designed by: Inderpreet Kaur Hayer   Group: Stanford BIOE44 - S11   (2015-10-24)
Revision as of 19:19, 26 October 2015 by Rna17 (Talk | contribs)

Arsenate Bioremediation Actuator in S. Cerevisiae


Assembly Compatibility:
  • 10
    INCOMPATIBLE WITH RFC[10]
    Illegal EcoRI site found at 2711
    Illegal XbaI site found at 1680
    Illegal PstI site found at 1035
  • 12
    INCOMPATIBLE WITH RFC[12]
    Illegal EcoRI site found at 2711
    Illegal PstI site found at 1035
  • 21
    INCOMPATIBLE WITH RFC[21]
    Illegal EcoRI site found at 2711
    Illegal BglII site found at 1471
  • 23
    INCOMPATIBLE WITH RFC[23]
    Illegal EcoRI site found at 2711
    Illegal XbaI site found at 1680
    Illegal PstI site found at 1035
  • 25
    INCOMPATIBLE WITH RFC[25]
    Illegal EcoRI site found at 2711
    Illegal XbaI site found at 1680
    Illegal PstI site found at 1035
  • 1000
    INCOMPATIBLE WITH RFC[1000]
    Illegal BsaI.rc site found at 2184


Design Notes

Below is an example of a possible implementation of BBa_M36532, the Arsenate Bioremediation Actuator. Due to size limitations, PHO84, ACR 1, and ACR 2, this actuator was created in two constructs, one with the PHO84 (M36533)[1] and another with ACR 1 and 2 (M36534) [2]. In this example, a GAL1 inducible sensor has been attached upstream for both designs. This image shows the different components of the Arsenate Bioremediation Actuator. Part numbers for the Kozak sequence and the PHO84, ACR 1, and ACR 2 genes can be found in the following section.


Example implementation:

http://imageshack.com/a/img907/2517/DITZFh.png


http://imageshack.com/a/img907/9338/NX12gL.png

Source

DNA 2.0 Plasmid (pD1201) with GAL1 inducible promoter, high copy number, and Leucine as the selectable marker. [3]

DNA 2.0 Plasmid (pD1204) with GAL1 inducible promoter, high copy number, and Histidine as the selectable marker. [4]

In order to have a multi-protein expression cassette, we incorporated an Internal Ribosome Entry Site (IRES), which was a T2A sequence. We got the amino acid sequence for this part from the following site [http://blog.addgene.org/plasmids-101-multicistronic-vectors]

Naturally occurring genes in S. cerevisiae: Pho84, ACR 1 and 2 Sequences were referenced from the yeast genome website and then were entered into the registry.

PHO84: BBa_M36535 [5]

ACR 1: BBa_M36536 [6]

ACR 2: BBa_M36537 [7]

References

Engineering of Saccharomyces cerevisiae for Bioremediation of Arsenate and Development of Yeast Vector Tools [http://gradworks.umi.com/35/02/3502760.htm]