*Promotions are valid in the US & Canada only. For international customers, please contact your local distributors. Promotion ends February 28, 2025. Discounts are not valid on previous purchases and cannot be combined with additional discounts.
Introduction
The GeneCopoeia EZShuttle™ cloning system uses the site-specific recombination machinery between the E. coli and phage lambda genomes for efficient DNA fragment transfer between plasmids, the same principle as Gateway® technology. The EZShuttle™ system includes the following major components. Click on each tab above for more details:
1. EZRecombinase™ Mixes for catalyzing attL x attR and attB x attP cloning reactions.
2. pEZ™ expression cloning vectors for creating ready-to-use expression clones.
3. EZShuttle™ Gateway® PLUS ORF clones. More than 35,000 human and mouse ORFs in pShuttle™ plasmid backbones.
The EZShuttle™ cloning system is fully compatible with Gateway® cloning, as shown in Table 1.

Table 1. Compatiblilty between components of GeneCopoeia’s EZShuttle™ recombination cloning system and Gateway® cloning technology.
Advantages
- Complete system of recombination cloning enzymes, expression cloning vectors, and sequence-verified ORF clones.
- Highly active for EZRecombinase™ LR Mix and EZRecombinase™ BP Mix.
- Choice of pEZ™ expression cloning vectors with a wide variety of selection markers and tags for cloning, processing, protein purification and gene function studies.
- Compatible with existing Gateway® cloning systems.
Applications
EZShuttle™ recombination cloning provides a highly fascile system for many powerful applications, including:
- Create ready-to-express expression clones. Readily shuttle ORFs or other DNA fragments of interest into an expression vector.
- Protein purification. Express your gene of interest in one or more systems for protein purification, leading to crystallization, antibody generation, DNA binding assays, and more.
- Promoter swapping. Transfer your gene of interest into vectors carrying different promoters.
- Promoter analysis. Test the function of multiple promoter candidates with a reporter such as luciferase.
- Gene tagging. Express your gene either untagged or with a multitude of in-frame fusion tags, such as 6XHis, Flag®, GFP, etc. Useful for co-immunoprecipitation, subcellular localization, etc.
Mechanism of EZShuttle™ cloning
GeneCopoeia’s EZShuttle™ recombination-based cloning system for DNA fragment transfer between plasmid vectors is based on the site-specific recombination machinery between the E. coli and phage lambda genomes, the same principle as Gateway® technology (Figure 1).

Figure 1. The attachment sites and proteins used for integration and excision reactions between E.coli and phage lambda genomes
Note: Gateway® is a registered trademark of Life Technologies.
BP & LR Mix
EZRecombinase™ BP Mix (Cat.No. ER003/ER004) Enhancement
GeneCopoeia’s EZRecombinase™ LR Mix and EZRecombinase™ BP Mix enable fast, efficient transfer of DNA fragments between compatible plasmids.The BP mix is used to create EZShuttle™ or Gateway® Entry clones by transferring fragments flanked by attB sites to attP-containing plamid vectors.
EZRecombinase™ BP II Enzyme Mix (Cat.No. ER013/ER014) Remix
Compared with BP Mix, BP enzyme and reaction buffer have been optimized to form a premixed solution, which is more stable and can be stored at -20°C.
Advantages
- Highly active.
- Compatible with existing Gateway® technology systems.
- 2 forms available: premixed and non-premixed.
EZRecombinase™ LR Mix (Cat.No. ER001/ER002) Enhancement
GeneCopoeia’s EZRecombinase™ LR Mix and EZRecombinase™ BP Mix enable fast, efficient transfer of DNA fragments between compatible plasmids. The EZRecombinase™ LR Mix is used to transfer fragments from EZShuttle™ or Gateway® Entry clones to pEZ™ or other Gateway®-compatible pDEST vectors.
Advantages
- Highly active. EZRecombinase™ LR Mix exhibits greater activity than Gateway® LR Clonase™.
- Compatible with existing Gateway® technology systems.
EZShuttle™ BP recombination technology
EZRecombinase™ BP mix contains Int and IHF, which catalyzes the in vitro recombination reaction between DNA fragments containing attB sites and vectors containing attP sites to generate new EZShuttle™ or Gateway® Entry clones.
A attB-PCR or Expression clone, a pShuttle™ vector, and EZRecombinase™ BP Mix are incubated at room temperature (25 ºC) for one hour and used to transform E. coli competent cells (Figure 2).

Figure 2. Transfer the DNA fragment containing the attB site to the pShuttle™ vector or Gateway® pDONR™ vector.
EZShuttle™ LR recombination technology
In LR recombination cloning, the transfer of a DNA fragment occurs between attL sites in an EZShuttle™ or Gateway® Entry clone and attR sites in a pEZ™ epression or Gatewayt® pDEST vector using EZRecombinase™ LR Mix, which contains Int (λ Integrase), IHF (E.coli Integration Host Factor), and Xis (Excisionase).
A pShuttle™ or Gateway® entry clone, a pEZ™ cloning destination vector, and EZRecombinase™ LR Mix are incubated at room temperature (25 ºC) for one hour and used to transform E. coli competent cells (Figure 3).

Figure 3. Principle for transfering a DNA fragment from an EZShuttle™ or Gateway® Entry clone to a pEZ™ expression or Gateway® pDEST vector using EZShuttle™ LR recombination cloning.
Shuttle clones
EZShuttle™ Gateway® PLUS ORF clones
GeneCopoeia offers more than 35,000 human and mouse EZShuttle™ Gateway® PLUS ORF clones. EZShuttle™ Gateway® PLUS ORF clones carry fully sequence-verified ORFs, and are available either with stop codons, to permit expression of ORFs that are either untagged or with in-frame N-terminal fusion tags, or without stop codons, allowing expression of ORFs with C-terminal fusion tags. ORFs are readily transferred to pEZ™ expression or Gateway® destination vectors using EZRecombinase™ LR Mix, as well as multiple cloning sites for conventional restriction enzyme-based cloning (Figure 3).
Click here for more information or to search the EZShuttle™ Gateway® PLUS ORF clone collections.
Advantages
- All ORFs are fully sequence verified.
- ORFs available with or without stop codons.
- Flanked by both attL sites and multiple restriction enzyme cloning sites for cloning flexibility.
- Can be used with pEZ™ expression or Gateway® destination vectors.

Figure 3. EZShuttle™ Gateway® PLUS ORF clones are flanked by sites for recombination-based cloning (attL1 and attL2), as well as multiple cloning sites (MC1 and MC2), and so can be used for both recombination as well as traditional restriction digestion ligation cloning.