Difference between revisions of "Part:BBa C0178"

Revision as of 14:16, 1 November 2017

autoinducer synthetase for PAI from Pseudomonas aeruginosa (no LVA)

same as C0078 except no LVA tag

Characterisation of LasI by Shanghaitech iGEM 2017

Group: Shanghaitech 2017

In synthetic biology, the quorum sensing system (QS system) has been used intensively for bacteria communication. The QS system has two parts: a generator of AHL molecules and a reporter which receives AHL molecules to activate downstream gene expression. For ‘Las’ QS system, the LasI protein catalyzes the enzymatic production of Las molecule (3OC12-HSL). A part expressing LasI has been deposited for more than a decade (since 2004), yet little information are available. This year this year we constructed two devices to ‘improve’ this part – ‘Las molecule generator BBa_K2315033 ’ and ‘Rpa-Las molecule converter BBa_K2315046’. The first one can generate Las molecules constitutively without any induction; the second one is a ‘converter’ which can receive another QS signal molecule Rpa and then produce Las. The mechanism of these two devices are shown below; the circuit design (.dna file) can be download in the part’s ‘design page’; the characterization of them can be found in this part’s ‘experiment page’. In conclusion, we successfully achieved the goal of ‘improve the function of an existing Biobrick Part’ with LasI coding sequence:

• 1) We verified LasI coding sequence is functional after more than a decade of deposition.
• 2) We used HPLC and LC-MS to quantitatively measure the product of LasI, Las (3OC12 HSL), which has not been done before.
• 3) We verified our two devices can produce Las successfully upon linking LasI to a constitutive promoter or a pRpa promotor induced with Rpa.
• 4) We verified that Las has a very long half-life, very stable in culture mixture.
• 5) We found Las concentration can be reported by real time GFP fluorescence upon mixing with a reporter bacteria when measured quantitatively using a plate reader.

Sequence and Features