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Team:The Agency Escondido/project
From 2013hs.igem.org
Project Description
Our project seeks to create two identical triads of transparent bacteria strains with complementary hydrophobic inducer production to emulate the game of "rock, paper, scissors" (Hydrophobic proteins can move through plasma membranes). Each triad will correspond to a group: group A and group B. Each type of bacteria will produce a specific protein that will act as an inducer to one and only one other bacteria strain(rock strain, paper strain, and scissors strain). Each triad will have different reporter genes designating its champion color. When two bacteria strains are combined in an agar plate, each protein secreted by the victorious strain, when acting as an inducer, changes the color of the cells on the plate identifying the champion.
Contents |
|---|
| Rock |
| Paper |
| Scissors |
Rock
The "rock" bacteria strain will have a CFP (part E0020) and promoter (part R0080). It will contain the LuxR gene (part C0062) to induce the "paper" strain.
Notebook
May 20
Andrew Buss began a protocol to prepare competent cells. This protocol omitted the final step of resuspension.
May 21
Andrew Buss completed the cell competency protocol from the previous day, spinning down 8 mL total in sequence.
With the arrival of ampicillin, Andrew Buss streaked BBa_S01022 onto a quadrant of an ampicillin agar plate.
Andrew Buss began transforming DNA for later assemblies into cells, using 5 uL of resuspended DNA per sample in water as suggested by the Parts Registry protocol. If the transformation failed with 5 uL, then 40 uL containing 1 ug could be used in another protocol. 50 pg of DNA from the transformation efficiency kit was used as an RFP control. The second and last centrifugation in the protocol was carried out using 0.6 mL tubes. Unfortunately, the rotor in the centrifuge was designed for 1.5 mL tubes. When centrifuging, three of the tubes were forced open, and fell through the holes in the rotor. These three samples were lost.
Andrew Buss restarted transformation, using the same protocol. Cells were left to incubate overnight in LB containing the appropriate antibiotics: the transformation control and pSB1C3 used chloramphenicol, while the remainder used ampicillin.
May 22
The chloramphenicol plate from the previous day displayed faint streaks (possibly simply indentations in the agar) but no distinct colonies. The ampicillin plate containing BBa_S01022 displayed dense growth. Andrew Buss extended some streaks onto the next quadrant to reduce the density to individual cells.
Andrew Buss streaked two quadrants of an ampicillin plate with BBa_R0080 and BBa_B0010. BBa_J37033 was transformed on the previous day, but was accidentally omitted from the streaking step.
Paper
The "paper" bacteria strain will have an GFP (part E0040) and promoter (part I1051). It will contain the Lambda cl repressor gene (part C0051) to induce the "scissors" strain.
Scissors
The "scissors" bacteria strain will have an RFP (part E1010)and promoter (part I12006). It will contain the AraC gene (part C0080) to induce the "rock" strain.
