(175a) Multiplex Recombineering-Enabled Genome Editing Tools

Recombineering has allowed researchers to rewire genomes much faster than traditional cloning techniques. With the advent of multiplex recombineering, genome-wide editing has become possible, allowing large scale reprogramming of cellular machinery. Despite these successes in large scale genome-wide reprogramming, the overall efficiency of recombineering still remains low, especially for changes > 6bp. Wang et al. have recently reported much higher efficiencies in recombineering by the use of co-selection, which enriches for cells that are highly recombinogenic¹. Therefore, we have designed a strain that has co-selection markers located around the genome and are easily screened/selected for, to enable high efficiency recombineering at any location. We have also developed a method to create large libraries of proteins with trackable, targeted changes in just a few days, further enabling genome editing and optimization. We will describe our efforts to apply these new tools for the rapid construction of protein, pathway, and combinatorial genome libraries.  

References

1              Wang, H. H. et al. Genome-scale promoter engineering by coselection MAGE. Nat Meth advance online publication, doi:http://www.nature.com/nmeth/journal/vaop/ncurrent/abs/nmeth.1971.html#supplementary-information (2012).