Part:BBa_K1010000

Ppnp ( 4-Nitrophenol responsive）

A promoter-included region from a 4-Nitrophenol(PNP) oxidation-related gene cluster. The promoter is consistently ON and specifically induced by PNP in the presence of NphR, a deduced regulator（Masahiro Takeo et al. 2008.）. We do a series of experiments about our sensor's transcription efficiency. We add PNP to liquid medium and incubated with our transgenic E.coli for several hours. And then we measure the fluorescence intensity with excitation wavelength at 470nm and emission wavelength at 509nm. We also measure absorption at 600nm which reflects E.coli's growth.

According to the experimental results, we find that our sensor does work. It shows that GFP transcription efficiency has a positive correlation relationship with time at the presence of PNP, though it will take a few hours before we can detect the changes. What's more, we think transcription efficiency may also have a positive correlation relationship with PNP concentration but at a very low level. The sensor will be in optimal situation when the concentration of PNP is 0.05mmol/L.

Electrophoretogram of nphR and Promoter nphA1

Efficiency of Promoter nphA1's Transcription with nphR

Cell Growth Curve with Promoter nphA1 and nphR

Acknowledgement

Thanks for Dr. Masahiro Takeo, who offered bacterium Rhodococcus sp. Strain PN1 and plasmids.

References

Takeo, Masahiro, Masumi Murakami, Sanae Niihara, Kenta Yamamoto, Munehiro Nishimura, Dai-ichiro Kato, and Seiji Negoro. "Mechanism of 4-nitrophenol oxidation in Rhodococcus sp. Strain PN1: characterization of the two-component 4-nitrophenol hydroxylase and regulation of its expression." Journal of bacteriology 190, no. 22 (2008): 7367-7374.

Yamamoto, Kenta, Munehiro Nishimura, Dai-ichiro Kato, Masahiro Takeo, and Seiji Negoro. "Identification and characterization of another 4-nitrophenol degradation gene cluster, nps, in Rhodococcus sp. strain PN1." Journal of bioscience and bioengineering 111, no. 6 (2011): 687-694.

Sequence and Features

Assembly Compatibility:

• 10
  COMPATIBLE WITH RFC[10]
• 12
  COMPATIBLE WITH RFC[12]
• 21
  COMPATIBLE WITH RFC[21]
• 23
  COMPATIBLE WITH RFC[23]
• 25
  COMPATIBLE WITH RFC[25]
• 1000
  INCOMPATIBLE WITH RFC[1000]

  Illegal BsaI.rc site found at 148