Endogenous Alkanes Induce

After systematic analysis of the microbial synthesis pathway from glucose to alkane in E.coli, we decide to choose the route from Fatty Acyl-ACP to alkane as the alkane producing module. Meanwhile, we conduct codon-optimization to the genes encoding NPDC and AAR based on the work done by Andreas Schirmer, making them more applicable to E.coli.

Here, we want to test the response of Alk-Sensor to endogenous alkanes. Construct:

We constructed 3 strains as shown in the figure below.

strain1:No.53：producing module (constitutive promoter J23100)+ Alk-Sensor device

strain2:No.54：producing module (constitutive promoter J23114) + Alk-Sensor device

strain3:No.55：Null+ Alk-Sensor device

The promoter strength of No.54 is stronger than that of No.53. Theorectically, alkane productivity of No.53 and No.54 will be different, and intracellular alkane concentration of the above 3 strains will be different-- No.54 the highest, No.53 lower and No.55 the lowest.

The 3 strains are cultured in 3ml M9 medium for about 24 hours.

Result：As shown in the diagram above, the fluorescent indensity of strain2 is the strongest, No.53 weaker and No.55 the weakest.

Conclusion:

(1) Alk-Sensor device can respond to the mixed intracellular alkanes, which can be used to detect alkanes inside the cell.

(2) There is a positive correlation between the concentration of intracellular alkanes, and the expression level of the reporter.