Part:BBa_K3190501
BAX: Bcl-2-associated X protein CDS
Coding sequence of the mammalian apoptosis regulator Bax protein. The gene will be integrated into the genome of the fungal chassis Saccharomyces cerevisiae, acting as the key gene in our biosafety device. Overexpression of Bax is lethal in S. cerevisiae, leading to the release of cytochrome c from mitochondria and thus apoptosis. The following results are characterization of a part submitted by iGEM17_NAU-CHINA: BBa_K2365048.
Usage and Biology
For our biosafety device, the Bax protein will be under the expression of a constitutive promoter, pADH2 (No part name specified with partinfo tag.). An anti-toxin, BAX Inhibitor-1 (BI-1) (BBa_K3190502) will be under the expression of an inducible promoter, pGAL1 (BBa_K3190050). As long as there is galactose present in the media, the anti-toxin will be expressed, inhibiting the apoptotic effects of Bax protein. If the cell escapes the media, the anti-toxin will no longer be expressed, and Bax protein will cause apoptosis of the cell.
Figure legend: Overview of our toxin/anti-toxin biosafety device.
Designing the construct
In order to characterize the Bax protein, we cloned it to inducible promoter pGAL1 (BBa_K3190050), as expression under a constitutive promoter would be expected to cause the cells to die.
Using USER ligation, we assembled the BAX gene with pGAL1 on a plasmid backbone compatible with multiplex integration cassette. The backbone used will integrate the construct in the yeast genome at chromosome 11, site ???.
E. coli cloning of pGAL1-BAX
Sequence and Features
- 10COMPATIBLE WITH RFC[10]
- 12COMPATIBLE WITH RFC[12]
- 21INCOMPATIBLE WITH RFC[21]Illegal BamHI site found at 452
- 23COMPATIBLE WITH RFC[23]
- 25COMPATIBLE WITH RFC[25]
- 1000COMPATIBLE WITH RFC[1000]
| None |