Part:BBa_K3081000
dCas9 binding with a degradation signal peptide ssrA
As it is found an big decrease in average nucleo-cytoplasmic ratio when treated with CRISPRri(BBa_K3081007)
targeted to OriC.In order to extend this system to other boxes which are shown to have over-inhibition on cell growth and small dynamic range, we improve the system to weaken its effect by adding a degradation signal peptide ssrA to dCas9. This largely accelerates the degradation rate of dCas9 and thus weaken its effect. Again, the CRISPRi system provides solid evidence for retention of dCas9 binding ability and degradation-promoting effect of ssrA. As a matter of fact, CRISPRi system with sgRNA targeted to mRFP coding region shows a more gentle decrease in fluorescence when dCas9 is fused with ssrA tag, while non-binding dCas9 with or without ssrA has no affect on mRFP expression. We tested the effect of the improved CRISPRri-ssrA system with target site to boxes which are shown to have excessive inhibition on cell growth, and found that the effect becomes much milder, which allows for a wider adjusting range.
Sequence and Features
- 10COMPATIBLE WITH RFC[10]
- 12COMPATIBLE WITH RFC[12]
- 21COMPATIBLE WITH RFC[21]
- 23COMPATIBLE WITH RFC[23]
- 25COMPATIBLE WITH RFC[25]
- 1000COMPATIBLE WITH RFC[1000]
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