Part:BBa_K4137012
CcdA antitoxin secretion pathway
The mleR regulatory site which comes from p_mles binds with the RBS and CcdA, the antitoxin that will bind with CcdB when induced by malate acid and detoxifies it, the purification tag 6xHis, and the terminator B1006 (BBa_B1006). When malate binds to the p_mleS promoter, it induces the production of CcdA, the antitoxin which assures chi18h8’s production around the plant roots.
Construct Design
We attached a Myc epitope Tag downstream of the ccdA sequence for purification purposes. A p_mleS promoter is attached upstream to the side of the open reading frame (ccdB-Myc coding complex) and RBS (AAGGAG). The terminator BBa_B1006 is attached downstream of the sequence.
Characterization
Cell harvest for ccdA was performed at 3h. We heated the mixture at 95˚C for 10 minutes, centrifuging at 4˚C, 16000g to extract the supernatant. An additional 95˚C heating and 100 µl PBS were added to eliminate clumping of pellets.
Sequence and Features
- 10COMPATIBLE WITH RFC[10]
- 12COMPATIBLE WITH RFC[12]
- 21COMPATIBLE WITH RFC[21]
- 23COMPATIBLE WITH RFC[23]
- 25COMPATIBLE WITH RFC[25]
- 1000COMPATIBLE WITH RFC[1000]
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