Bacterial Artificial Chromosome Mutagenesis Using Recombineering

Table 1

Comparing conventional BAC modification strategies with recombineering.

	Conventional strategies	Recombineering
BAC Modification Strategies	Homologous recombination: RecBCD and RecF pathways	RecE pathway: RecE, RecT (+Gam)
	Site-specific integration: Integrases of different pathways (Cre, Flp)	Lambda Red pathway: Exo, Beta, Gam
Flexibility	Require pre-engineered or existing specific sites: Chi sites, recombinase attachment sites	Sequence independent, but need prior knowledge of target sequence
Homology requirement	Long homology of ~1 kb is required for efficient homologous recombination	As short as 40 bp of homology is sufficient
Versatility	May require specific strain backgrounds to work	Mobile systems can be easily transferred to and adapted for use in a wide range of strains and species
Efficiency and facileness	Low efficiency for homologous recombination	At least 50-fold higher than traditional homologous recombination
	Labourious and lengthy	Can be done in days