Difference between revisions of "Part:BBa K2273113:Design"

 
 
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Sites of restriction enzymes generating compatible overhangs are indicated by sharing one color. (<span style="color:blue">EcoRI</span> and <span style="color:blue">PstI</span> are marked in blue, <span style="color:green">NotI</span> in green, <span style="color:red">XbaI</span> and <span style="color:red">SpeI</span> in red
 
Sites of restriction enzymes generating compatible overhangs are indicated by sharing one color. (<span style="color:blue">EcoRI</span> and <span style="color:blue">PstI</span> are marked in blue, <span style="color:green">NotI</span> in green, <span style="color:red">XbaI</span> and <span style="color:red">SpeI</span> in red
  
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As the overall promoter sequence was amplified from the <i>B.subtilis</i> W168 genome using primers featuring the RFC10 standard restriction sites:
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The promoter sequence was amplified from the <i>B.subtilis</i> W168 genome using primers featuring the RFC10 standard restriction sites:
  
 
Forward Primer Sequence containing RFC10 prefix: 5`-gatcGGAATTCGCGGCCGCTTCTAGAAATCACAATTGATAAAGCTTTCTAA-3`
 
Forward Primer Sequence containing RFC10 prefix: 5`-gatcGGAATTCGCGGCCGCTTCTAGAAATCACAATTGATAAAGCTTTCTAA-3`
  
−
Revers Primer Sequence containing RFC10 suffix: 5`-GATCCTGCAGCGGCCGCTACTAGTTCAAATGATTTTGTATTACCTTTG-3`
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Revers Primer Sequence containing RFC10 suffix: 5`-gatcCTGCAGCGGCCGCTACTAGTTCAAATGATTTTGTATTACCTTTG-3`
  
  
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===References===
 
===References===
 +
[http://www.subtiwiki.uni-goettingen.de/v3/gene/view/713BAB7190E1F86C55103049B29072F00E0DFFB3]
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[http://www.uniprot.org/uniprot/P39824]
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 +
Marta Toth <i>et al.</i> (2015) Class D Beta-Lactamases Do Exist in Gram-Positive Bacteria. <i>Nature Chemical Biology</i> <b>(12)</b>: 9-14.

Latest revision as of 13:21, 3 October 2017


Short version of the Promoter PpenP found in B. subtilis


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]


Design Notes

This Promoter was designed to be cloned in front of the luxABCDE operon serving a a reporter gene in the pBS3C vector backbone. Therefore, the RFC10 Standard Restriction sites have been added as prefix and suffix via PCR.

RFC10 prefix and suffix sequences:

Prefix with EcoRI, NotI, XbaI GAATTCGCGGCCGCTTCTAGA
Suffix with SpeI, NotI and PstI ACTAGTAGCGGCCGCTGCAGA

Sites of restriction enzymes generating compatible overhangs are indicated by sharing one color. (EcoRI and PstI are marked in blue, NotI in green, XbaI and SpeI in red

The promoter sequence was amplified from the B.subtilis W168 genome using primers featuring the RFC10 standard restriction sites:

Forward Primer Sequence containing RFC10 prefix: 5`-gatcGGAATTCGCGGCCGCTTCTAGAAATCACAATTGATAAAGCTTTCTAA-3`

Revers Primer Sequence containing RFC10 suffix: 5`-gatcCTGCAGCGGCCGCTACTAGTTCAAATGATTTTGTATTACCTTTG-3`



Source

This part sequence was derived from the Bacillus subtilis W168 genome sequence.

References

[http://www.subtiwiki.uni-goettingen.de/v3/gene/view/713BAB7190E1F86C55103049B29072F00E0DFFB3]

[http://www.uniprot.org/uniprot/P39824]

Marta Toth et al. (2015) Class D Beta-Lactamases Do Exist in Gram-Positive Bacteria. Nature Chemical Biology (12): 9-14.