Part:BBa_K5300016

nifH promoter, a kind of promoter responding to oxygen content

Promoter responsive to low oxygen signals, activates downstream gene transcription under low oxygen conditions and becomes inactive in high oxygen environment.

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
  COMPATIBLE WITH RFC[1000]

In order to verify the function of nifH promoter, we constructed a composite circuit of nifH promoter and gfp, which was seamlessly cloned into pBBR1MCS-2 along with “glnK promoter+sgRNA-gfp” for subsequent experiments. (Figure 2-1-4).

Fig. 2-1-4 The model of promoter validation.

Since Cas12k was not expressed and the glnK promoter was repressed by high-nitrogen conditions, we cultured rhizobia with this plasmid in high-nitrogen (1 g/L NH4Cl) M9 medium. Under these conditions it was thought that the glnK promoter would be unable to express subsequent genes and that gfp expression would not be repressed. The fluorescence intensity was detected after 24 h of incubation under both low and high oxygen conditions, respectively (Figure 2-1-5).

Fig. 2-1-5 Ratio of relative fluorescence intensity to OD value of bacterial solution. Student's t-test, ns: no significant difference; *, p-value < 0.05; **, p-value<0.01; ***, p-value < 0.001.

Apparently, the expression intensity of nifH promoter was up-regulated under low-oxygen conditions; while under high-oxygen conditions, the expression intensity of nifH promoter was down-regulated. It was successfully verified that the nifH promoter was repressed by hyperoxic conditions.