Regulatory
Pnif

Part:BBa_K1796001

Designed by: Nannan Xie   Group: iGEM15_SCU_China   (2015-09-17)
Revision as of 19:06, 18 September 2015 by Nianhui (Talk | contribs) (/*  A                                        )

promoter of Paenibacillus sp. WLY78 nif gene cluster

The promoter of Paenibacillus sp. WLY78 nif gene cluster, We named it Pnif. Pnif is a σ70-dependent promoter located upstream of nifB. Pnif is in control of all the gene clusters of nitrogen fixation genome.


Usage and Biology

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]


Confirmation of nif promoter

We standardized the nif promoter of Paenibacillus sp. WLY78, and linked it to the upstream of 3 reporter genes (RFP reporter: BBa_K1357010, YFP reporter: BBa_E0430;amilCP reporter: BBa_K1357009) to confirm the reliability of this promoter. The results are shown in Figure 2.

Confirmation_of_Pnif_AB.png

 A                                                                                        B

622px-C-confirmation_of_Pnif.jpeg

  C

Fig.4. Confirmation of nif promoter of Paenibacillus sp. WLY78. (A) Pnif+RFP reporter; (B) Pnif+YFP reporter;(C)Pnif+amilCP reporter.

The results confirmed the reliability of Pnif. The initiation of Pnif does not require inducement. Thus it is a effective constitutive promoter.


Functional Parameters

Flow cytometry test

We also tested Pnif expression with flow cytometry. A GFP biobrick part BBa_E0240 was linked downstream of this promoter, then we tested both Pnif sample and a blank control sample simultaneously. The GFP expression by Pnif was clearly shown in the result

434px-Flow_cytometry_test.jpeg

All the data of blank control was shown in the left, and the data of CFP expression was shown in the right.

800px-Flow_cytometry_test2.png

Relative promoter strength estimates

Reference Strength
BBa_K1796001 13
BBa_K176011 67
typeconstitutive promoter

SCU_China's MEASUREMENT

800px-Comparison_of_Pnif_and_Pd.jpeg

[edit]
Categories
//chassis/prokaryote/bcepacia
//direction/forward
//regulation
//regulation/constitutive
//rnap/prokaryote/ecoli/sigma70
Parameters
typeconstitutive promoter