Part:BBa_K3081046
pBAD-dCas9-T7-sgRNA
Unlike traditional CRISPRi system, our CRISPRri systemBBa_K3081060 is driven by an arabinose-inducible promoter with low leakage and great switch property. The expression of sgRNA is driven by a novel inducible T7 system. But the sgRNA is a relatively meaningless sequence. And two BsaI sites flanking the meaningless sequence allow for easy assembly of intended sgRNA sequence by Golden Gate method. After Golden Gate assembly, the ‘sgRNA’ sequence will be replaced by our carefully selected sequences (like M+, BBa_K3081107; R1+,BBa_K3081108; etc.). These newly generated pBAD-dCAS9-T7-M+/R1/… parts each contains a meaningful sgRNA which guides the dCas9 protein to different boxes on E.coli genome replication initiation region, OriC, thus blocking the binding of relevant proteins essential for replication initiation to their boxes.
Sequence and Features
- 10COMPATIBLE WITH RFC[10]
- 12INCOMPATIBLE WITH RFC[12]Illegal NheI site found at 1205
Illegal NheI site found at 2321
Illegal NheI site found at 6708 - 21INCOMPATIBLE WITH RFC[21]Illegal BamHI site found at 1144
Illegal BamHI site found at 4600
Illegal XhoI site found at 6437 - 23COMPATIBLE WITH RFC[23]
- 25INCOMPATIBLE WITH RFC[25]Illegal AgeI site found at 979
- 1000INCOMPATIBLE WITH RFC[1000]Illegal BsaI site found at 6627
Illegal BsaI.rc site found at 6223
Illegal SapI site found at 961
Illegal SapI.rc site found at 5369
| None |