DNA

Part:BBa_K2100050:Design

Designed by: Trinh Nguyen   Group: iGEM16_MIT   (2016-10-19)


pENTR L4_EGSH 2x k-turn_R1


Assembly Compatibility:
  • 10
    INCOMPATIBLE WITH RFC[10]
    Illegal EcoRI site found at 1287
    Illegal XbaI site found at 2193
    Illegal PstI site found at 1780
    Illegal PstI site found at 2200
  • 12
    INCOMPATIBLE WITH RFC[12]
    Illegal EcoRI site found at 1287
    Illegal NheI site found at 832
    Illegal NheI site found at 1098
    Illegal PstI site found at 1780
    Illegal PstI site found at 2200
    Illegal NotI site found at 2207
  • 21
    INCOMPATIBLE WITH RFC[21]
    Illegal EcoRI site found at 1287
    Illegal BglII site found at 1410
  • 23
    INCOMPATIBLE WITH RFC[23]
    Illegal EcoRI site found at 1287
    Illegal XbaI site found at 2193
    Illegal PstI site found at 1780
    Illegal PstI site found at 2200
  • 25
    INCOMPATIBLE WITH RFC[25]
    Illegal EcoRI site found at 1287
    Illegal XbaI site found at 2193
    Illegal PstI site found at 1780
    Illegal PstI site found at 2200
  • 1000
    INCOMPATIBLE WITH RFC[1000]
    Illegal SapI.rc site found at 709


Design Notes

The distance from the k-turn repeats to the open reading frame of the downstream gene determines the efficiency of L7Ae/k-turn repressing system. Thus, we were considering adding k-turns right after the promoter (in the pENTR promoter vector) or right in front of the gene coding sequence (in the pENTR gene vector).


Source

EGSH was obtained using PCR extension from pENTR pEGSH (BBa_K2100021). The k-turn sequence was obtained from addgene (plasmids deposite by Dr. Ron Weiss' lab). We ordered the k-turns as single-stranded oligos with different Q sites for Golden Gate reaction.

References