Part:BBa_K1020005
Alkane Marker--Device for detection of various alkanes
AlkR is a transcriptional regulator belonging to AraC/Xyls family, which could detect a broad range of alkanes and alkenes with carbon chain length from C7 to C36. It is said that AlkR is the only bioreporter that is able to detect alkane with carbon chain length greater than C18. The mechanism is that Alkane were presumably recognized by AlkR regulatory protein and their interaction triggered a conformation change of AlkR dimers which isomerizes the promoter-RNAP complex and lead to the activation of alkM gene. AlkR regulatory protein includes C-terminal DNA-binding domain(CTD) for promoter binding and N-terminal domain(NTD) for inducer recognition. Here, different from BBa_K1020004,we use a weaker consititutive promoter.
We use different kinds of long chain alkane to characterize the part.
Then we test Octane(C8),Decane(C10),Dodecane(C12), Tetradecane(C14), Hexadecane(C16) and Octadecyl(C18) respectively.
Strains are cultured in M9 medium for about 12 hours. Adjust the concentration of suspension culture until OD reaches 0.6, then take out 3mL and add 300μL. After that, induce for 24 hours.
Measure the ratio of fluorescent indensity (fluorescent indensity per OD) of cells, which is shown below:
From this diagram, we can clearly see that induced by saturated Octane(C8), ratio of fluorescent indensity is the strongest.
Then we choose C8 and C14 for upper limits of detection.Because C8 is commonly used to characterize alkane-sensor in most references, and Pentadecane(C15) is our endogenous alkane producing module produce this kind of alkane.
Sequence and Features
- 10COMPATIBLE WITH RFC[10]
- 12INCOMPATIBLE WITH RFC[12]Illegal NheI site found at 1028
Illegal NheI site found at 1051 - 21COMPATIBLE WITH RFC[21]
- 23COMPATIBLE WITH RFC[23]
- 25INCOMPATIBLE WITH RFC[25]Illegal AgeI site found at 752
Illegal AgeI site found at 864 - 1000INCOMPATIBLE WITH RFC[1000]Illegal BsaI.rc site found at 1821
| None |