Difference between revisions of "Part:BBa K1362000:Design"
Line 6: Line 6:
  
 
===Design Notes===
 
===Design Notes===
−
Between the coding sequences of the ''Npu'' DnaE C-intein and the N-intein we placed <partinfo>BBa_J04450</partinfo>, an mRFP selection marker flanked by BsaI sites that can be replaced by a protein of interest. A strong RBS <partinfo>BBa_K1362090</partinfo> was added.  
+
Between the coding sequences of the ''Npu'' DnaE C-intein <partinfo>BBa_K1362401</partinfo>and the N-intein <partinfo>BBa_K1362400</partinfo>we placed <partinfo>BBa_J04450</partinfo>, an mRFP selection marker flanked by BsaI sites that can be replaced by a protein of interest. A strong RBS <partinfo>BBa_K1362090</partinfo> was added.  
 
This part was assembled by CPEC [[#References|[1]]] from PCR products of <partinfo>BBa_K1362093</partinfo> , <partinfo>BBa_J04450</partinfo>, pVS07 and pVS41 [[#References|[2]]].  
 
This part was assembled by CPEC [[#References|[1]]] from PCR products of <partinfo>BBa_K1362093</partinfo> , <partinfo>BBa_J04450</partinfo>, pVS07 and pVS41 [[#References|[2]]].  
  

Revision as of 13:06, 17 October 2014

NpuDnaE intein RFC[105] circularization construct


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    COMPATIBLE WITH RFC[12]
  • 21
    COMPATIBLE WITH RFC[21]
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    INCOMPATIBLE WITH RFC[25]
    Illegal AgeI site found at 931
    Illegal AgeI site found at 1043
  • 1000
    INCOMPATIBLE WITH RFC[1000]
    Illegal BsaI site found at 1220
    Illegal BsaI.rc site found at 145


Design Notes

Between the coding sequences of the Npu DnaE C-intein BBa_K1362401and the N-intein BBa_K1362400we placed BBa_J04450, an mRFP selection marker flanked by BsaI sites that can be replaced by a protein of interest. A strong RBS BBa_K1362090 was added. This part was assembled by CPEC [1] from PCR products of BBa_K1362093 , BBa_J04450, pVS07 and pVS41 [2].


Source

Assembled from subparts: ...


References

[1] Quan, J. & Tian, J. Circular polymerase extension cloning for high-throughput cloning of complex and combinatorial DNA libraries. Nat. Protoc. 6, 242–51 (2011).

[2] Zettler, J., Schütz, V. & Mootz, H. D. The naturally split Npu DnaE intein exhibits an extraordinarily high rate in the protein trans-splicing reaction. FEBS Lett. 583, 909–14 (2009).