Parts used to characterize and build our final project
Component | Description | Part/Accession # | Base Pairs | Plasmid | Resistance | Well |
RBS | Ribosomal Binding Site | BBa_B0034 | 12 | pSB1A2 | Ampicillin | plate 1, 2M |
Cph8 | Red Light sensor | BBa_I15010 | 2,238 | pSB2K3 | Kanamycin | N/A |
RFP | Red Fluorescent Protein | BBa_J04051 | 720 | N/A | ||
OmpR (E. coli) | description | NP_417864.1 | 720 | pSB1T3 | Tetracycline | N/A |
OmpR (R. sphaeroides) | description | sequence | 720 | plasmid | resistance | N/A |
Terminator | Stops Transcription | BBa_B0015 | 129 | pSB1AK3 | Ampicillin and Kanamycin | plate 1, 23L |
OmpR + Terminator | description | sequence | 916 | pany-amp | Ampicillin | synthesized |
OmpC promoter | description | BBa_R0082 | 108 | pSB1A2 | Ampicillin | plate 1, 16K |
puc B/A | description | sequence | 375 | pSB1A3 | Ampicillin | N/A |
puc B | description | YP_353390 | 156 | ? | ? | N/A |
puc A | description | YP_353391 | 165 | ? | ? | N/A |
OmpC promoter+BA | description | sequence | 539 | pany-kana | Kanamycin | synthesized |
Light Response System | description | BBa_M30109 | 4,333 | ? | Ampicillin | N/A |
TetR repressible | description | BBa_J13002 | 74 | pSB1A2 | Ampicillin | plate 1, 13B |
Green Fluorescent Protein | Marker for characterization | BBa_E0240 | 876 | pSB1A2 | Ampicillin | plate 1, 12M |
Plasmid | Description | Base Pairs | Resistance | Copy Number |
pSB1A2 | BioBrick Vector | 2,079 | Ampicillin | high |
pSB1K3 | 2,206 | Kanamycin | high | |
pSB1A3 | Assembly plasmid | 2,157 | Ampicillin | high |
pSB2K3 | 4,425 | Kanamycin | variable | |
pSB1T3 | Assembly plasmid | 2,463 | Tetracycline | high |
pSB1AK3 | Assembly Plasmid | 3,189 | Ampicillin and Kanamycin | high |
pANY | Synthesized from Geneart | |||
pRKCBC3 | Tetracycline |
Component | Description | Part/Accession # | Base Pairs | Plasmid | Resistance |
RBS+Cph8 | description | BBa_K227000 | Base Pairs | plasmid | Resistance |
RBS+Cph8+RBS+OmpR+Terminator | description | BBa_K227001 | Base Pairs | plasmid | Resistance |
Cph8 (resubmission) | description | BBa_K227002 | Base Pairs | plasmid | Resistance |
RBS+Cph8+RBS+OmpR+Terminator +OmpC promoter+pucBA | description | BBa_K227003 | Base Pairs | plasmid | Resistance |
puc A | description | BBa_K227004 | Base Pairs | plasmid | Resistance |
puc B | description | BBa_K227005 | Base Pairs | plasmid | Resistance |
puc BA | description | BBa_K227006 | Base Pairs | plasmid | Resistance |
puc promoter | description | BBa_K227007 | Base Pairs | plasmid | Resistance |
OmpC+GFP | description | BBa_K227008 | Base Pairs | plasmid | Resistance |
RBS+Cph8+OmpC+GFP | description | BBa_K227009 | Base Pairs | plasmid | Resistance |
OmpR (optimized for Rhodobacter Sphaeroides) | description | BBa_K227010 | Base Pairs | plasmid | Resistance |
whole construct | description | BBa_K227011 | Base Pairs | plasmid | Resistance |
The first part of our characterization begins with the puc promoter. The puc promoter is what turns on the entire system naturally in Rhodobacter sphaeroides. The puc promoter is what ultimately controls the number of LH2 light harvesting complexes, which is how our system will yield an increase in photosynthetic efficiency. It is important that we are able to compare the transcription rate of the puc promoter in the two systems so that we can determine exactly how much efficiency is gained by adding a red light sensor. By attaching Green Fluorescent Protein (GFP) to the promoter we can quantify the rate of transcription by measuring the emission of green light using a fluorescence spectrophotometer. We would expect to see more fluorescence with more transcription and vis versa.
Modeling the Gene Regulatory Network
References
1. Alon, Uri. Introduction to systems biology and the design principles of biological networks. Boca Raton, FL: Chapman & Hall, 2006.
2. Bower, James M. Computational Modeling of Genetic and Biochemical Networks (Computational Molecular Biology). New York: M.I.T. PRESS, 2001.
3. System modeling in cellular biology from concepts to nuts and bolts. Cambridge, MA: MIT P, 2006.