Difference between revisions of "Part:BBa K3081032"
| Line 7: | Line 7: | ||
https://2019.igem.org/wiki/images/4/49/T--Peking--R1%2B%2820bp%29_ssrA.gif | https://2019.igem.org/wiki/images/4/49/T--Peking--R1%2B%2820bp%29_ssrA.gif | ||
| − | R1+20bp | + | R1+ssrA 20bp |
https://2019.igem.org/wiki/images/6/6b/T--Peking--R1%2B%2819bp%29_ssrA.gif | https://2019.igem.org/wiki/images/6/6b/T--Peking--R1%2B%2819bp%29_ssrA.gif | ||
| − | R1+19bp | + | R1+ssrA 19bp |
https://2019.igem.org/wiki/images/0/05/T--Peking--R1%2B%2818bp%29_ssrA.gif | https://2019.igem.org/wiki/images/0/05/T--Peking--R1%2B%2818bp%29_ssrA.gif | ||
| − | R1+18bp | + | R1+ssrA 18bp |
Revision as of 18:14, 21 October 2019
pBAD-dCas9-ssrA-J23119-R1+(18bp)
This composite part is the principal design of the inducible CRISPR-based DNA replication interference system, with the 18 bp sgRNA targeting to the R1+ DnaA box on E.coli genome replication initiation region, OriC. In natural situations, R1+ is a high affinity box for DnaA binding. By blocking the binding of DnaA protein to R1+ box using a 18bp sgRNA, alleviation of severe arrest and inhibition to the genome replication initiation is achieved. This part is an improvement of BBa_K3081009, which we add a degradation signal peptide ssrA to the dCas9. This largely accelerates the degradation rate of dCas9 and weakens its overinhibtion on genome replication initiation targeted to the R1 box.
R1+ssrA 20bp
R1+ssrA 19bp
R1+ssrA 18bp
Sequence and Features
- 10COMPATIBLE WITH RFC[10]
- 12INCOMPATIBLE WITH RFC[12]Illegal NheI site found at 1205
Illegal NheI site found at 5459
Illegal NheI site found at 5482 - 21INCOMPATIBLE WITH RFC[21]Illegal BglII site found at 1470
Illegal BamHI site found at 1144 - 23COMPATIBLE WITH RFC[23]
- 25INCOMPATIBLE WITH RFC[25]Illegal AgeI site found at 979
- 1000INCOMPATIBLE WITH RFC[1000]Illegal SapI site found at 961