Difference between revisions of "Part:BBa K3081041"

Latest revision as of 19:19, 21 October 2019

CRISPR-based replication interference system for p15A plasmid copy number control, ini(-) site

This composite part uses a CRISPR-based DNA replication interference system for the copy number control of the p15A plasmid. The dCas9 is expressed in an inducible manner with a ssrA degradation tag fused to its C-terminal to minimize the basal expression. The sgRNA has a 20-bp complementary sequence with the top strand (complementary to the strand that replication activator (RNA II) binds to) of initiation site (+1) on p15A origin. It can efficiently downregulate the plasmid copy number but has some unexpected leackage.

Design of the p15A plasmid copy number control system

The effect of 4 target sites on p15A origin

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.

Sequence and Features

Assembly Compatibility:

10
COMPATIBLE WITH RFC[10]
12
INCOMPATIBLE WITH RFC[12]
Illegal NheI site found at 1205
Illegal NheI site found at 2321
Illegal NheI site found at 5495
Illegal NheI site found at 5518
21
INCOMPATIBLE WITH RFC[21]
Illegal BamHI site found at 1144
Illegal BamHI site found at 4600
23
COMPATIBLE WITH RFC[23]
25
INCOMPATIBLE WITH RFC[25]
Illegal NgoMIV site found at 5522
Illegal AgeI site found at 979
1000
INCOMPATIBLE WITH RFC[1000]
Illegal SapI site found at 961

@@ Line 15: / Line 15: @@
 <b>The effect of 4 target sites on p15A origin</b>
 </center>
+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.
 <!-- Add more about the biology of this part here