Difference between revisions of "Part:BBa K3081046"

Latest revision as of 19:21, 21 October 2019

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

Reference:

[1]Wolański M, Donczew R, Zawilakpawlik A, et al. oriC-encoded instructions for the initiation of bacterial chromosome replication.[J]. Frontiers in Microbiology, 2015, 5(735):735.