In-fusion assembly: seamless engineering of multidomain fusion proteins, modular vectors, and mutations.
In-Fusion can join any two pieces of DNA that have a 15-bp overlap at their ends. The result is equivalent to a recombination event at the ends of the DNAs. The 15-bp overlap may be engineered by inclusion in primers used to PCR amplify a segment of DNA. Originally described for inserting one piece of DNA into a restriction enzyme-digested plasmid, we have found In-Fusion can join four or more pieces of DNA in a single reaction. We used this insight to construct seamless fusion proteins, modular vectors with readily interchangeable segments, and novel mutagenesis strategies. Replacement In-Fusion can be used to delete any desired DNA segment in a plasmid and replace it with any desired new DNA segment without limitations on position or size.
Zhu
Baogong
B
Dana-Farber Cancer Institute, Harvard Medical School, Boston, MA 02115, USA.
Cai
Guifang
G
Hall
Emily O
EO
Freeman
Gordon J
GJ
D004247:DNA / Q000235:genetics*; D004252:DNA Mutational Analysis; D005818:Genetic Engineering / Q000379:methods*; D005822:Genetic Vectors / Q000235:genetics*; D009154:Mutation; D017434:Protein Structure, Tertiary; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000235:genetics*; D017422:Sequence Analysis, DNA / Q000379:methods*; D012984:Software
D011993:Recombinant Fusion Proteins; D004247:DNA