Part:BBa_K2457006
Cas9 device + Optimized sgRNA
This BioBrick is composed by BBa_K2457000 (AraC + pBAD), BBa_K2457001 (Cas9) and BBa_K2457002 (simple guide RNA). Which AraC + pBAD regules the Cas9 expression through your repressor and your inductor: Glucose and L-Arabinose, respectively. Cas9 is a protein that with the assistance of sgRNA cleaves a target-site in the bacterial genome. This construction was developed by Amazonas_Brazil team to be a fundamental part of the CRISPeasy toolbox, divided in CRISPReasy-out and CRISPReasy-in.
CRISPeasy-out is a standardized knock out approach that aims to turn the current method in an easier way to engineer the bacterial genome by Non-Homologous End Joining (NHEJ).
CRISPeasy-in is a standardized knock-in approach that aims to turn the current method in an easier way to insert a donor template into the bacterial genome by Homology-Directed Repair (HDR).
Figure 1: BBa K2457006 circuit
Usage and Biology
Design
Characterization
Sequencing
Figure 2:Sequencing electropherogram from BBa_K2457006.
Figure 3:Alignment of the designed sequence and our final construction from BBa_K2457006.
Sequence and Features
- 10COMPATIBLE WITH RFC[10]
- 12COMPATIBLE WITH RFC[12]
- 21INCOMPATIBLE WITH RFC[21]Illegal BamHI site found at 1207
- 23COMPATIBLE WITH RFC[23]
- 25INCOMPATIBLE WITH RFC[25]Illegal AgeI site found at 1042
- 1000INCOMPATIBLE WITH RFC[1000]Illegal SapI site found at 1024
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