Difference between revisions of "Part:BBa K1796001"

(Flow cytometry test)
(SCU_China's MEASUREMENT)
Line 53: Line 53:
 
====SCU_China's MEASUREMENT====
 
====SCU_China's MEASUREMENT====
 
https://static.igem.org/mediawiki/2015/thumb/9/98/Comparison_of_Pnif_and_Pd.jpeg/800px-Comparison_of_Pnif_and_Pd.jpeg
 
https://static.igem.org/mediawiki/2015/thumb/9/98/Comparison_of_Pnif_and_Pd.jpeg/800px-Comparison_of_Pnif_and_Pd.jpeg
 +
<p>Comparison of the expression efficiency of Pnif and Pd. Pd: Promoter BBa_J23100; Pnif: nif promoter.</p>

Revision as of 19:32, 18 September 2015

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

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

Comparison of the expression efficiency of Pnif and Pd. Pd: Promoter BBa_J23100; Pnif: nif promoter.