Difference between revisions of "Part:BBa K3365065"

Latest revision as of 13:18, 27 October 2020

Fourth lure sequence upstream of eGFP

This part is an off-target detecting platform for dCas9-ω. We put this part in the plasmid and transform it into △rpoZ-MG1655, a strain of E.coli that is koncked out rpoZ. If dCas9-ω is binding to the lure sequence, the subnit Omega fused to dCas9 C-terminal can activate the expression of reporter by recruiting the RNAP holoenzyme and the cell would emit green fluorescence,which means dCas9-ωis off-target, else the weak promoter and the lack of rpoZ makes the reporter cannot express or just have a quite low expression level.

According to the prediction model, the off-target rate of the lure sequence in this part is 0.37543%.

Fig:Schematic diagram of this part

Sequence and Features

Assembly Compatibility:

10
COMPATIBLE WITH RFC[10]
12
INCOMPATIBLE WITH RFC[12]
Illegal NheI site found at 66
Illegal NheI site found at 89
21
INCOMPATIBLE WITH RFC[21]
Illegal BamHI site found at 834
Illegal XhoI site found at 843
23
COMPATIBLE WITH RFC[23]
25
COMPATIBLE WITH RFC[25]
1000
COMPATIBLE WITH RFC[1000]

@@ Line 3: / Line 3: @@
 <partinfo>BBa_K3365065 short</partinfo>
-Fourth lure sequence upstream of eGFP
+This part is an off-target detecting platform for dCas9-ω. We put this part in the plasmid and transform it into △<i>rpoZ</i>-MG1655, a strain of E.coli that is koncked out <i>rpoZ</i>. If dCas9-ω is binding to the lure sequence, the subnit Omega fused to dCas9 C-terminal can activate the expression of reporter by recruiting the RNAP holoenzyme and the cell would emit green fluorescence,which means dCas9-ωis off-target, else the weak promoter and the lack of <i>rpoZ</i> makes the reporter cannot express or just have a quite low expression level.
+According to the prediction model, the off-target rate of the lure sequence in this part is 0.37543%.
+[[File:Lure_sequence_upstream_of_eGFP.png]]
+<b>Fig:</b>Schematic diagram of this part
 <!-- Add more about the biology of this part here