Difference between revisions of "Part:BBa K5300016"
 
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<partinfo>BBa_K5300016 parameters</partinfo>
 
<partinfo>BBa_K5300016 parameters</partinfo>
 
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         <p>In order to verify the function of <i>nifH</i> promoter, we constructed a composite circuit of nifH promoter and <i>gfp</i>, which was seamlessly cloned into pBBR1MCS-2 along with "<i>glnK</i> promoter+sgRNA-GFP" for subsequent experiments. (Figure 1).</p>
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         <p>In order to verify the function of <i>nifH</i> promoter, we constructed a composite circuit of <i>nifH</i> promoter and <i>gfp</i>, which was seamlessly cloned into pBBR1MCS-2 along with “<i><i>glnK</i></i> promoter+sgRNA-<i>gfp</i>” for subsequent experiments. (Figure 2-1-4).</p>
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         <figcaption><b>Fig. 1 The model of <i>nifH</i> promoter validation.</b>> Student's t-test, ns: no significant difference; *, p-value < 0.05; **, p-value<0.01; ***, p-value < 0.001.</figcaption>
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         <figcaption><b>Fig. 2-1-4 The model of promoter validation.</b></figcaption>
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        <p>Since <i>Cas12k</i> was not expressed and the <i><i>glnK</i></i> 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 <i><i>glnK</i></i> promoter would be unable to express subsequent genes and that <i>gfp</i> 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).</p>
  
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            Since Cas12k was not expressed and the <i>glnK</i> 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 <i>glnK</i> promoter would be unable to express subsequent genes and that <i>gfp</i> 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).
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         <figcaption><b>Fig. 2-1-5 Ratio of relative fluorescence intensity to OD value of bacterial solution.</b> Student's t-test, ns: no significant difference; *, p-value < 0.05; **, p-value<0.01; ***, p-value < 0.001.</figcaption>
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         <p>Apparently, the expression intensity of <i>nifH</i> promoter was up-regulated under low-oxygen conditions; while under high-oxygen conditions, the expression intensity of <i>nifH</i> promoter was down-regulated. It was successfully verified that the <i>nifH</i> promoter was repressed by hyperoxic conditions.</p>
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         <figcaption><b>Fig. 2 Ratio of relative fluorescence intensity to OD value of bacterial.</b> Student's t-test, ns: no significant difference; *, p-value < 0.05; **, p-value<0.01; ***, p-value < 0.001.</figcaption>
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            Apparently, the expression intensity of <i>nifH</i> promoter was up-regulated under low-oxygen conditions; while under high-oxygen conditions, the expression intensity of <i>nifH</i> promoter was down-regulated. It was successfully verified that the <i>nifH</i> promoter was repressed by hyperoxic conditions.
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Latest revision as of 21:27, 1 October 2024


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.