This biobrick has a promoter inducible by PAI+LasR (BBa_R0079)and produces LuxI enzyme (BBa_F1610). This enzyme produces 3OC6HSL (AI).
The DNA of BBa_R0079 and BBa_F1610 biobricks where used.
BBa_K091146 promoter (PAI+LasR inducible & AI+LuxR repressible) controls the expresion of LasI enzyme, no terminator
The biobricks BBa_K091146 and BBa_S03154 where used to build this biobrick
LasR/LasI Inducible & RHLR/RHLI repressible Promoter
This part is similar to BBa_R0079 but has an added binding site for RHLR/RHLI in between the -35 and -10 region. Thus this promoter is induced by LasR/LasI but repressed by RHLR/RHLI binding.
The natural spacer between the -35 and -10 is 14bp and the natural O1 site for RHLR/RHLI is 20bp long. There was not much over lap of the -35 or the -10 with this sequence, so it was nessicary to chop off some of the binding site sequence in order to maintain the correct spacing and preserve the -35 and -10 sequence. The binding site sequence is missing a tcc on is 5' end and a t on its 3' end.
Rust L. et. al., "Analysis of the Pseudomonas aeruginosa Elastase (lasB) Regulatory Region". Journal of Bacteriology, 1996.
Complex Quorum sensing circuit that receives the signal of PAI+LasR and AI+LuxR to control the production of LuxI and LasI enzymes.
The biobricks BBa_K266006 and BBa_K266000 where used to build this biobrick
K091146 is a modified version of the pLas' promoter (BBa_K091117) containing the addition of a luxR binding site between the -35 and -10 regions. This promoter is expected to be activated in the presence of PAI-1 and LasR and repressed in the presence of AI-1 and LuxR.
The Lux box was placed adjacent to the LasR binding site with the aim of minimizing cross-talk in the presence of both PAI-1 and AI-1; since the LasR site should have a higher affinity, we hope that the binding of PAI-1 will competitively exclude the binding of AI-1.
The LuxR binding site is from BBa_R0062 and the pLas' promoter is from the region upstream of the LasI gene in Pseudomonas aeruginosa.
Vikram Vijayan, Allen Hsu, Lawrence Fomundam
LasR and 3OC12HSL regulated promoter with a single Tet Operator (binding site for TetR repressor)
LasR and 3OC12HSL regulated promoter with CI Operator 1 (binding site for lambda cI repressor)
this hybrid promoter is designed to be consecutively off.It combines with plasR(R0079) and lambda cI OR1(see C0051).Adding P-AI-1 sould activate the promoter, while repressed by lambda cI.
"CI binding of the OR region. It binds OR1 with highest affinity, OR2 with next highest, and OR3 with lowest"(http://www.nonanone.com/qw/files/Biology%204450/test2_lecture8,9.txt)
PAI+LasR -> LasI+GFP & AI+LuxR --| LasI+GFP
BBa_K091146 promoter (PAI+LasR inducible & AI+LuxR repressible) controls the policistronic expresion of LasI enzyme and GFP, double terminator
The biobricks BBa_K266005 and BBa_E0840 where used to build this biobrick.
This pLas' promoter contains the regulatory region upstream of the LasI gene in Pseudomonas aeruginosa. K091117 includes a LasR binding site as well as a -10 region modified from CATAAA to TATAAA to reflect our pLasXOR promoter design. Activation is expected to occur in the presence of the autoinducer PAI-1 and the LasR protein. The pLas' promoter serves as a control for the evaluation of the pLasXOR promoter, part BBa_K091118.
To minimize leaky transcription, a secondary transcriptional start site was excluded from the design.
The sequence of the LasI upstream regulatory region is available at http://www.pubmedcentral.nih.gov/articlerender.fcgi?artid=176640
K091157 is a modified version of the pLux' promoter (BBa_K091156) containing the addition of a LasR binding site between the -35 and -10 regions. This promoter is expected to be activated in the presence of AI-1 and LuxR and repressed in the presence of PAI-1 and LasR.
In order to fit the LasR binding site between the -35 and -10 regions, the last base of the Las Box was cropped and the -35 region was modified to overlap with the first two bases of the Las Box.
The LasR binding site is from the region upstream of the LasI gene in Pseudomonas aeruginosa. The rest of the promoter is the same as BBa_K091156.
Binding region for LasR protein (positive regulation)
"Analysis of the Pseudomonas aeruginosa Elastase (lasB) Regulatory Region". Lynn Rust, Everett Pesci, and Barbara Iglewski