Difference between revisions of "Part:BBa K191003:Design"
(→‎Design Notes)
 
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* Notice : 2 restriction sites of PstI in LovTAP sequence so LovTAP can't be cut with PstI.
 
* Notice : 2 restriction sites of PstI in LovTAP sequence so LovTAP can't be cut with PstI.
−
 
  
 
* LovTAP was obtained by PCR and ligated into double-term plasmid ( LovTAP was cut with E and S, plasmid E and X)
 
* LovTAP was obtained by PCR and ligated into double-term plasmid ( LovTAP was cut with E and S, plasmid E and X)
−
* LOVTAP-Term is then amplified by PCR and cut with E and X to be put into another plasmid of high-copy and Kanamycine resistance.
+
* LovTAP-Term is then amplified by PCR and cut with E and X to be put into another plasmid of high-copy and only Kanamycine resistance.
 
* LacI was ligated into RBS
 
* LacI was ligated into RBS
−
* PCR on LacI-RBS, digestion with E and S then ligated into LovTAP-Term plasmid which was cut by E and X.  
+
* PCR on LacI-RBS, digestion with E and S then ligated into LovTAP-Term plasmid which was cut by E and X.
  
 +
===Source===
  
−
===Source===
+
<partinfo>BBa_R0010</partinfo>, <partinfo>BBa_B0030</partinfo>, <partinfo>BBa_B0015</partinfo>.
  
−
BBa_R0010, BBa_B0030, BBa_B0015.
 
 
Plasmid containing LovTAP gene was kindly provided by the authors of : Light-activated DNA binding in a designed allosteric protein; Strickland et al., Proceedings of the National Academy of Sciences (2008) vol. 105 (31) pp. 10709
 
Plasmid containing LovTAP gene was kindly provided by the authors of : Light-activated DNA binding in a designed allosteric protein; Strickland et al., Proceedings of the National Academy of Sciences (2008) vol. 105 (31) pp. 10709
  

Latest revision as of 13:28, 21 October 2009

Promoter Lac I - RBS - LovTAP - Term


Assembly Compatibility:
  • 10
    INCOMPATIBLE WITH RFC[10]
    Illegal PstI site found at 370
    Illegal PstI site found at 627
  • 12
    INCOMPATIBLE WITH RFC[12]
    Illegal PstI site found at 370
    Illegal PstI site found at 627
  • 21
    COMPATIBLE WITH RFC[21]
  • 23
    INCOMPATIBLE WITH RFC[23]
    Illegal PstI site found at 370
    Illegal PstI site found at 627
  • 25
    INCOMPATIBLE WITH RFC[25]
    Illegal PstI site found at 370
    Illegal PstI site found at 627
  • 1000
    INCOMPATIBLE WITH RFC[1000]
    Illegal SapI.rc site found at 764


Design Notes

  • Notice : 2 restriction sites of PstI in LovTAP sequence so LovTAP can't be cut with PstI.
  • LovTAP was obtained by PCR and ligated into double-term plasmid ( LovTAP was cut with E and S, plasmid E and X)
  • LovTAP-Term is then amplified by PCR and cut with E and X to be put into another plasmid of high-copy and only Kanamycine resistance.
  • LacI was ligated into RBS
  • PCR on LacI-RBS, digestion with E and S then ligated into LovTAP-Term plasmid which was cut by E and X.

Source

BBa_R0010, BBa_B0030, BBa_B0015.

Plasmid containing LovTAP gene was kindly provided by the authors of : Light-activated DNA binding in a designed allosteric protein; Strickland et al., Proceedings of the National Academy of Sciences (2008) vol. 105 (31) pp. 10709

References

Light-activated DNA binding in a designed allosteric protein; Strickland et al., Proceedings of the National Academy of Sciences (2008) vol. 105 (31) pp. 10709