Difference between revisions of "Part:BBa K4115043"

Revision as of 06:45, 5 October 2022

lac UV5 promoter-nifA-PnifH-sfGFP-B0015

This composite part is used to regulate PnifH by nifA protein. In the original A.caulinodans genome, the expression of the downstream PnifH can make nitrogen-fixing gene clusters functional. Thus， we transformed this composite part into the backbone of the pBBR1 shuttle plasmid and transferred it into A.caulinodans ORS571 to verify whether the nifA protein could regulate PnifH by observing the response of the downstream sfGFP protein.

Usage and Biology

The nifA gene is regulated by the lac UV5 promoter. When there is no IPTG induction, the lac UV5 promoter is inhibited and the nifA cannot be expressed. When induced by IPTG, the lac UV5 promoter is activated to express the downstream nifA. The expressed nifA protein then regulates PnifH to activate the downstream sfGFP expression. (Figure 1)
Thus, we can conclude whether the nifA-PnifH regulating system is functional by observing the fluorescence intensity of sfGFP.

Figure 1: Gene pathway map of the nifA-PnifH regulating system.

Sequence and Features

Reference

[1] Ryu, MH., Zhang, J., Toth, T. et al. Control of nitrogen fixation in bacteria that associate with cereals. Nat Microbiol 5, 314–330 (2020). doi: https://doi.org/10.1038/s41564-019-0631-2