Difference between revisions of "Part:BBa K1796001"
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<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 σ70-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 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. |
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<!-- 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> | |
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<span class='h3bb'>Sequence and Features</span> | <span class='h3bb'>Sequence and Features</span> | ||
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| − | <!-- 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 | ||
| + | === A B=== | ||
| + | https://static.igem.org/mediawiki/2015/thumb/3/34/C-confirmation_of_Pnif.jpeg/622px-C-confirmation_of_Pnif.jpeg | ||
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| + | === C=== | ||
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| + | <p>Confirmation of nif promoter of Paenibacillus sp. WLY78. (A) Pnif+RFP reporter; (B) Pnif+YFP reporter;(C)Pnif+amilCP reporter.</p> | ||
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| + | 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> | + | ====Flow cytometry test==== |
| − | <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> |
| − | + | 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 | ||
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| + | ====Relative promoter strength estimates==== | ||
| + | |||
| + | {|border="1" | ||
| + | !Reference | ||
| + | !Strength | ||
| + | |- | ||
| + | |<partinfo>BBa_K1796002</partinfo> | ||
| + | |13 | ||
| + | |- | ||
| + | |<partinfo>BBa_K176011</partinfo> | ||
| + | |67 | ||
| + | |} | ||
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<partinfo>BBa_K1796001 parameters</partinfo> | <partinfo>BBa_K1796001 parameters</partinfo> | ||
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| + | ====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
- 10COMPATIBLE WITH RFC[10]
- 12COMPATIBLE WITH RFC[12]
- 21COMPATIBLE WITH RFC[21]
- 23COMPATIBLE WITH RFC[23]
- 25COMPATIBLE WITH RFC[25]
- 1000COMPATIBLE 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.
A B
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
All the data of blank control was shown in the left, and the data of CFP expression was shown in the right.
Relative promoter strength estimates
| Reference | Strength |
|---|---|
| BBa_K1796002 | 13 |
| BBa_K176011 | 67 |
| type | constitutive promoter |
SCU_China's MEASUREMENT
Comparison of the expression efficiency of Pnif and Pd. Pd: Promoter BBa_J23100; Pnif: nif promoter.