Difference between revisions of "Part:BBa K191005:Design"

 
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__NOTOC__
 
__NOTOC__
 
<partinfo>BBa_K191005 short</partinfo>
 
<partinfo>BBa_K191005 short</partinfo>
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===Design Notes===
 
===Design Notes===
−
1.5 step- PCR was used.
 
  
 +
* This biobrick was created using our new method, which we named [http://2009.igem.org/wiki/index.php?title=Team:EPF-Lausanne/Protocols/1.5_steps_PCR protocol 1.5-step PCR].
 +
* Long primers are added first to amplify the DNA from the plasmid (this sequence is a part of <partinfo>BBa_I6007</partinfo>). Then short primers with iGEM prefix and suffix are added to amplify the entire sequence.
 +
 +
1st Forward primer: 5'-gtttcttcgaattcgcggccgcttctagagtggcaaatattctgaaatgagctgttgacaattaatcatcgaactagttaactagtacgcaagttcacgtaaaaagggtatcgacaaagaggagaaatactagatgtcc-3'
 +
 +
2nd Forward primer: 5'-gtttcttcgaattcgcggccgcttctagagtggcaaatattctgaaatgagctgttgacaattaatcatcgaactagttaactagta-3'
 +
 +
1st Reverse primer: 5'-cagctcctcgcccttgctcaccatctagtatttctcctctttctctagtagtgc-3'
 +
 +
2nd Reverse primer: 5'-ctagctagcctctagtagtgctcagtatctctatcac-3'
 +
 +
* Trp promoter till TetR promoter was constructed with 1.5 step PCR then cut with E and S to be ligated into iGEM plasmid with RFP ( which was cut with E and X).
  
  
 
===Source===
 
===Source===
  
−
BBa_I13507 for RFP and BBa_I6007 for TetR
+
<partinfo>BBa_I13507</partinfo> for RFP.
 +
 
 +
<partinfo>BBa_I6007</partinfo> for TetR.
 +
 
 +
<partingo>BBa_K191007</partinfo>
  
 
===References===
 
===References===

Revision as of 17:09, 21 October 2009

TRP promoter - RBS - TetR - Term - TetR promoter - RBS - RFP - Term


Assembly Compatibility:
  • 10
    INCOMPATIBLE WITH RFC[10]
    Illegal SpeI site found at 43
    Illegal SpeI site found at 51
  • 12
    INCOMPATIBLE WITH RFC[12]
    Illegal SpeI site found at 43
    Illegal SpeI site found at 51
  • 21
    COMPATIBLE WITH RFC[21]
  • 23
    INCOMPATIBLE WITH RFC[23]
    Illegal SpeI site found at 43
    Illegal SpeI site found at 51
  • 25
    INCOMPATIBLE WITH RFC[25]
    Illegal SpeI site found at 43
    Illegal SpeI site found at 51
    Illegal AgeI site found at 1576
    Illegal AgeI site found at 1688
  • 1000
    COMPATIBLE WITH RFC[1000]


Design Notes

  • This biobrick was created using our new method, which we named [http://2009.igem.org/wiki/index.php?title=Team:EPF-Lausanne/Protocols/1.5_steps_PCR protocol 1.5-step PCR].
  • Long primers are added first to amplify the DNA from the plasmid (this sequence is a part of BBa_I6007). Then short primers with iGEM prefix and suffix are added to amplify the entire sequence.

1st Forward primer: 5'-gtttcttcgaattcgcggccgcttctagagtggcaaatattctgaaatgagctgttgacaattaatcatcgaactagttaactagtacgcaagttcacgtaaaaagggtatcgacaaagaggagaaatactagatgtcc-3'

2nd Forward primer: 5'-gtttcttcgaattcgcggccgcttctagagtggcaaatattctgaaatgagctgttgacaattaatcatcgaactagttaactagta-3'

1st Reverse primer: 5'-cagctcctcgcccttgctcaccatctagtatttctcctctttctctagtagtgc-3'

2nd Reverse primer: 5'-ctagctagcctctagtagtgctcagtatctctatcac-3'

  • Trp promoter till TetR promoter was constructed with 1.5 step PCR then cut with E and S to be ligated into iGEM plasmid with RFP ( which was cut with E and X).


Source

BBa_I13507 for RFP.

BBa_I6007 for TetR.

<partingo>BBa_K191007</partinfo>

References