Experiment

Endogenous Induce&Resistance Selection

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Here, we want to test the resistance against tetracycline of Alk-Sensor induced by endogenous alkanes.

We add alkanes outside the cell to test the function of TetA:

Construct:

Left: No.60: TetA downstream of Palkm and AlkR downstream of J23103, are cloned into vector PSB1C3 and there’s a alkane producing vector PSB3K3.

Right: No.61: We construct a blank vector PSB3K3 without NPDC and AAR as blank control, and the other vector PSB1C3 is the same as No.60.

Characterization:

Strains of No.60 are cultured in LB medium for about 12 hours. Add 17μg/mL Kan, 12μg/mL Cm and 12μg/mL Tc respectively. After that, inoculate into 3 ml LB medium for an overnight cultures at 37 ℃ with 220rpm shaking.

Result:

Alkane producing E.coli can grow in tube but blank vector E.coli cannot grow .

Conclusion:

Synthesized alkanes induce the expression of alkR, which enables E.coli to survive under the pressure of Tetracycline.

Thus, we can get a 3×3 matrix, with promoter strength from weak, medium to strong, and plasmid copy number from small, medium to large.