Part:BBa_K5127012
Genome Integration of butyrate production protein (pReplace-Tes4)
The pReplace construction is designed for the integration of GOI into the E.coli MG1655 genome, with GFP downstream serving as a reporter gene to visualize successful integration. The two red features in the figure represent LP-2 homologous regions, which are 500bp sites on the genome utilized for integrase λ-Red mediated plasmid integration. Additionally, stringent ORI: R6K is incorporated, requiring a specialized protein for replication that most bacteria do not produce. This ensures that upon entering the host cell, the plasmid either cannot replicate and is eliminated or becomes integrated into the chromosome and replicates alongside it. This approach guarantees successful plasmid integration into the genome.
Sequence and Features
- 10INCOMPATIBLE WITH RFC[10]Illegal PstI site found at 501
- 12INCOMPATIBLE WITH RFC[12]Illegal NheI site found at 129
Illegal NheI site found at 152
Illegal PstI site found at 501 - 21INCOMPATIBLE WITH RFC[21]Illegal BglII site found at 530
- 23INCOMPATIBLE WITH RFC[23]Illegal PstI site found at 501
- 25INCOMPATIBLE WITH RFC[25]Illegal PstI site found at 501
Illegal AgeI site found at 923 - 1000COMPATIBLE WITH RFC[1000]
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