Difference between revisions of "Part:BBa K1796001"

 
(13 intermediate revisions by one other user not shown)
Line 2: Line 2:
 
<partinfo>BBa_K1796001 short</partinfo>
 
<partinfo>BBa_K1796001 short</partinfo>
  
The promoter of Paenibacillus sp. WLY78 nif gene cluster, We named it Pnif. Pnif is a &#963;70-dependent promoter located upstream of nifB. Pnif is in control of all the gene clusters of nitrogen fixation genome.  
+
The promoter of Paenibacillus sp. WLY78 nif gene cluster, We named it Pnif. Pnif is a &#963;70-dependent promoter located upstream of nifB. Pnif is in control of all the gene clusters of nitrogen fixation genome. It is a constitutive promoter,which can successfully function in E.coli.
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.
+
A                      B                    C
+
  
 
 
https://static.igem.org/mediawiki/2015/b/b2/Confirmation_of_Pnif_AB.png
 
https://static.igem.org/mediawiki/2015/thumb/3/34/C-confirmation_of_Pnif.jpeg/622px-C-confirmation_of_Pnif.jpeg
 
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.
+
<!-- Add more about the biology of this part here-->
 
+
 
+
<!-- Add more about the biology of this part here
+
 
===Usage and Biology===
 
===Usage and Biology===
 
+
<p></p>
 
<!-- -->
 
<!-- -->
 
<span class='h3bb'>Sequence and Features</span>
 
<span class='h3bb'>Sequence and Features</span>
Line 23: Line 13:
  
  
<!-- Uncomment this to enable Functional Parameter display  
+
<!-- Uncomment this to enable Functional Parameter display-->
 +
===Confirmation of nif promoter===
 +
<p>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.</p>
 +
https://static.igem.org/mediawiki/2015/b/b2/Confirmation_of_Pnif_AB.png
 +
===&nbsp;A&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;B===
 +
https://static.igem.org/mediawiki/2015/thumb/3/34/C-confirmation_of_Pnif.jpeg/622px-C-confirmation_of_Pnif.jpeg
 +
 
 +
===              &nbsp;    C===
 +
 
 +
 
 +
 
 +
<p>Confirmation of nif promoter of Paenibacillus sp. WLY78. (A) Pnif+RFP reporter; (B) Pnif+YFP reporter;(C)Pnif+amilCP reporter.</p>
 +
 
 +
The results confirmed the reliability of Pnif. The initiation of Pnif does not require inducement. Thus it is a effective constitutive promoter.
 +
 
 
===Functional Parameters===
 
===Functional Parameters===
<p>Relative promoter strength estimates</p>
+
====Flow cytometry test====
<p>123456123456</p>
+
<p>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</p>
2222222222222222
+
https://static.igem.org/mediawiki/2015/thumb/e/e0/Flow_cytometry_test.jpeg/434px-Flow_cytometry_test.jpeg
 +
<p>All the data of blank control was shown in the left, and the data of CFP expression was shown in the right.</p>
 +
https://static.igem.org/mediawiki/2015/thumb/6/62/Flow_cytometry_test2.png/800px-Flow_cytometry_test2.png
 +
 
 +
====Relative promoter strength estimates====
 +
 
 +
{|border="1"
 +
!Reference
 +
!Strength
 +
|-
 +
|<partinfo>BBa_K1796002</partinfo>
 +
|13
 +
|-
 +
|<partinfo>BBa_K176011</partinfo>
 +
|67
 +
|}
 +
 
 
<partinfo>BBa_K1796001 parameters</partinfo>
 
<partinfo>BBa_K1796001 parameters</partinfo>
 
<!-- -->
 
<!-- -->
 +
 +
====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
 +
<p>Comparison of the expression efficiency of Pnif and Pd. Pd: Promoter BBa_J23100; Pnif: nif promoter.</p>

Latest revision as of 20:45, 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. It is a constitutive promoter,which can successfully function in E.coli.


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_K1796002 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.