This device accepts PoPS as input and produces the LuxI enzyme. This enzyme produces 3OC6HSL.
IPTG inducible AHL generator, LasI device.
pConstitutive: J23100 LasI device: K772005
Sender device which accepts PoPS input and produces an enzyme CinI. This enzyme produces the 3OC14HSL signaling molecule.
3OC6HSL Sender controlled by pBad
Debra, Fred, Jay, Madeleine, Cell-Cell Signaling SMUG
The pBad promoter controlling a LuxI sender device. The protein is an untagged LuxI enzyme which produces an AHL (3OC6HSL).
The pictures below are from a time course, where pRCV-3 produced GFP via HSL-bound LuxR promotion. I13015 was on the left half of the slide (induced in .1% arabinose, thereby producing the HSL for pRCV-3), while pRCV-3 was on the right half. Controls were done with pRCV-3 alone, I13015 alone and the two together without any arabinose. In all three cases little or no GFP was expressed.
Sensory rhodopsin I (SRI) with HtrII & Tar, sensitive to orange light
Sensory rhodopsin I fusion with HtrII & Tar, which is sensitive to orange light, with sensing spectra covering from 590-650 nm. We used it for sensing blur-green light to achieve phototaxis and switch for downstream gene expression.
Pfam (version 26.0) was used to predict the domain of fusion protein. Linker was refereed and modified from previous research study of SRI fusion with HtrII.
From genomic sequence of bacterial N. pharaonis (DSM 2160) and E.coli K12.
LacI regulated ethylene generator that uses 3 enzymes, blah
This is a coding sequence of S. cerevisiae mating pheromone together with a secretion tag. Expression in the yeast cell causes secretion of the mature mating pheromone into the medium.
In the genome, the secretion tag is followed by a three copies of the coding sequence for the actual peptide. We had to amplify only the secretion tag first, and then added the DNA sequence of the peptide using different reverse primer.
Obtained from the genome of S. cerevisiae [MF(ALPHA)1 locus].
Quorum sensing is the mechanism of communication between bacteria via synthesizing and detecting chemical compounds in the media. With these mediators, the promoters and their yield can be tuned or regulated. This part represents an alternative quorum sensing sender device which is designed to induce PoPS production in the parts including pLas such as BBa_K772004. LasI is an enzyme generator gene sequences which synthesizes 6HSLO3 (AHL) in order to induce a PoPS response in the pLas promoter. pLas promoter works in the presence of both 6HSLO3 and LasR protein.
Note that there is an RBS sequence just before LasI; in order to be easily used with quorum sensing receiver devices such as BBa_F2620 or BBa_K772004.
LasI sequence is gathered from Registry.
Sensory rhodopsin II (SRII) with HtrII & Tsr, sensitive to blue-green light
Sensory rhodopsin II fusion with HtrII & Tsr, which is sensitive to blue-green light, with sensing spectra covering from 400-500 nm. We used it for sensing blur-green light to achieve phototaxis and switch for downstream gene expression.
Pfam (version 26.0) was used to predict the domain of fusion protein. Linker was refereed and modified from previous research study of SRII fusion with HtrII.
Red light triggering phototaxis using chp8
Red light triggering phototaxis using chp8, which can sense the red light from 600-700 nm.
6 poly-His tag was positioned at C-terminal of the protein.
The sequence of chp8 is from previous biobrick.
This BioBrick contains a ''virA'' receptor from ''Agrobacterium tumefaciens'' C58 under the control of a constitutive promotor. We used it to mutate ''virA'' by error-prone PCR. For screening the mutants BBa_K389011 is also needed.
The ''virA'' gene is under the control a medium strong constitutive promotor.
This part generates the TaZ receptor. The TaZ is under a Pcat constitutive promoter. TaZ is a fused Tar-EnvZ receptor meant for Aspartate dependent response, by activating the EnvZ/OmpR two component signalling system and thus activating the PompC promoter. We used this part for prove-of-concept for low concentration detection. By using this TaZ Generator we were able to test different concentrations of Aspartate and their effect on the PompC promoter present in another plasmid.
We had to add a gggggg sequance between RBS and TaZ sequances.
Pcat: BBa_I14033 RBS: BBa_B0034 TaZ: BBa_C0082 dTer: BBa_B0015
Simple inducible (LacI) ethylene generator
A simpler version of BBa_K417006 which expresses ACC synthase and ACC oxidase (EFE) to produce ethylene. This version does not express the SAM synthetase enzyme (BBa_K417000) and relies on S-adenosyl methionine generated by the chassis.
Native LacI promoter (BBa_R0010) was chosen since it facilitates easier assembly compared with the shorter alternative BBa_R0011, although it would be worth testing both these promoters. It is unknown if the order of the coding sequences is important in this device.
See part documentation for enzyme coding sequences BBa_K417001 and BBa_K417002 for details on the origin of the sequences.
High Population->mRFP (pLac->LuxI; pLacIQ->luxR; pLux->mRFP;)
Quorum sensing circuit under a pLac promoter. Population turns red (mRFP) once quorum sensing kicks in.
Double terminator not used after LuxI because it is unnecessary and has the potential to make the plasmid more unstable.
Built through assembly of BBa_K084014 and BBa_J10000.
a luxI homologous AHL synthetase from Sinorhizobium meliloti 1021
The Sinorhizobium meliloti strain Rm1021 harbors at least two quorum-sensing systems; one of them is sinR/sinI system, The sinI is responsible for the production of several novel AHLs, including C12-HSL, 3-Oxo-C14-HSL, C16:1-HSL, 3-Oxo-C16:1-HSL, C18-HSL, some of theses are the longest AHLs identified so far. The sin system correlates with a delay in the appearance of nitrogen-fixing nodules, as well as with an overall decrease in the number of pink nodules, suggesting a role for quorum sensing in establishing a successful symbiosis with Medicago sativa. As a autoinducer synthatase, the use of sinI is the same with luxI, namely to produce AHLS.
The Sinorhizobium meliloti 1021 is a mode organism in Rhizobia. so We got the S. meliloti 1021 from Nanjing Agriculture University easily, and then cloned the sinI gene from S. meliloti 1021 with primers designed by the part design tool in the website ginkgobioworks.com.
SinI is from the Sinorhizobium meliloti 1021. We get its gene sequence from the www.microbesonline.org, and then cloned them through PCR with biobrick part primers.
This protein generator produces the enzyme LuxI (no ssrA tag). This enzyme metabolized the small intracellular molecule, SAM, to produce 3OC6HSL.
Sensory rhodopsin II (SRII) with HtrII & Tar, sensitive to blue-green light
Sensory rhodopsin II fusion with HtrII & Tar, which is sensitive to blue-green light, with sensing spectra covering from 400-500 nm. We used it for sensing blur-green light to achieve phototaxis and switch for downstream gene expression.
Sender device which accepts PoPS input and produces an enzyme LasI. This enzyme produces the AI-1 signaling molecule
Sender device which accepts PoPS input and produces an enzyme RhlI. This enzyme produces the N-butyryl-HSL signaling molecule.