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Figure 2 | Microbial Cell Factories

Figure 2

From: Knock-in/Knock-out (KIKO) vectors for rapid integration of large DNA sequences, including whole metabolic pathways, onto the Escherichia coli chromosome at well-characterised loci

Figure 2

Generic features of the KIKO plasmids and integration protocol. A) Map of the general features of the KIKO vectors, showing the location of the antibiotic resistance cassettes (ApR, ampicillan resistance; KmR, kanamycin resistance; CmR, chloramphenicol resistance), the flippase recombinase target sites (FRT), the hairpin loops (HL), the homologous arms (HA1 and HA2) used for recombination onto the E. coli chromosome and the unique restriction sites located within the multiple cloning site (MCS). B) Process for the isolation of integration mutants. This represents the fastest possible generation protocol by standard cloning; in practice, strain construction is often delayed be a few days for quality control (QC). Strain construction can be shortened by two days if insertion cassettes are constructed by Gibson assembly.

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Microbial Cell Factories

ISSN: 1475-2859

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