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FAQ: How large a DNA fragment can I assemble?

Gibson Assembly Cloning Kit (containing NEB 5-alpha Competent E. coli ) has been used to successfully clone a 15 kb DNA fragment into a 5.4 kb plasmid in E. coli, totaling up to 20.4 kb in length. However, as a general rule for assembled products greater than 10 kb, NEB recommends NEB 10-beta Competent E. coli (High Efficiency, NEB #C3019) or NEB 10-beta Electrocompetent E. coli (NEB #C3020).