Recombinational Cloning

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Recombinational cloning became popular with the introduction of three cloning systems: Gateway®, Creator™, and Echo Cloning™ systems. These systems use a site-specific recombinase (Integrase in Gateway and Cre Recombinase in Creator and Echo) to allow reliable transfer of a fragment from one vector to another without using restriction enzymes and ligases. 

Typically, a researcher would clone a sequence of interest into a holding vector ("Entry" for Gateway and "Donor" for Creator) using traditional cloning methods. Once the new clone is made, it is easily shuttled to many different "destination" or "acceptor" vectors that contain the appropriate sequence recognized by the recombinase (attachment sites attB and attP with Gateway and loxP with Creator/Echo). Higher throughput is possible with these systems and they have become a useful tool for screening many different expression hosts for protein expression projects or for multiple reporter vectors for functional analysis studies. At this time, only the Gateway system is still commercially supported, although NEB does sell Cre Recombinase (NEB #M0298), an essential reagent for the in vitro recombination step used by the Creator and Echo Cloning systems.

Recombinational Cloning Workflow

Note that times are based on estimates for moving a gene from one plasmid to another. If the source for gene transfer is gDNA, add 2 hours to calculation for the traditional cloning method. Total time does not include transformation, isolation or analysis.

Featured Products:
Monarch™ Plasmid Miniprep Kit
Monarch™ DNA Gel Extraction Kit
Monarch™ PCR & DNA Cleanup Kit (5 μg)
Cre Recombinase
NEB 5-alpha Competent E.coli
Q5® Hot Start High-Fidelity DNA Polymerase

Find Additional Products for use in Recombinational Cloning applications:
DNA Preparation
Restriction Enzyme Digestion
Reverse Transcription (cDNA Synthesis)
DNA Isolation
DNA Analysis
Restriction Enzyme Digestion
Colony PCR
DNA Sequencing