Mediating the recombination reaction
As explained, the core of Invitrogen™ Gateway™ cloning is the Entry vector. Once the Entry clone is ready, the gene of interest is easily shuttled to a secondary plasmid, the Destination vector. This reaction is mediated by a robust enzyme mixture called LR Clonase™, which contains the necessary protein activity to excise the gene of interest from the Entry clone and integrate it into the Destination vector, which then becomes your expression clone (Figure 1). Reversing this reaction simply entails performing a BP reaction with BP Clonase enzyme mix. Both LR and BP Clonase enzyme mixtures are easy-to-use master mix formats ensuring consistency and reliability from reaction to reaction.
Types of Clonase enzymes
Gateway BP Clonase enzyme contains both Int (Integrase) and IHF (Integration Host Factor) proteins that catalyze the in vitro recombination of PCR products or DNA segments from clones (containing attB sites) and a Donor vector (containing attP sites) to generate Entry clones. View the selection guide below to find the best BP Clonase™ enzyme for your research goals.
Gateway LR Clonase II enzyme mix contains a proprietary blend of Int (Integrase), IHF (Integration Host Factor) and Xis (Excisionase) enzymes that catalyze the in vitro recombination between an Entry clone (containing a gene of interest flanked by attL sites) and a Destination vector (containing attR sites) to generate your expression clone. We offer different formats of LR Clonase enzyme mixtures, depending on your application and desired format. View the selection guide below to find the best LR Clonase enzyme for your research goals.
Figure 1. The Gateway reactions.
Clonase Enzyme Selection Guide
|LR Clonase Enzyme Mixes|
|Best||LR Clonase II Plus Enzyme Mix||Single mix of enzyme and reaction buffer|
|Very good||LR Clonase II Enzyme Mix||Single mix of enzyme and reaction buffer|
|Good||LR Clonase Enzyme||Enzyme|
5X reaction buffer
|BP Clonase Enzyme Mixes|
|Best||BP Clonase II Enzyme Mix||Single mix of enzyme and reaction buffer|
|Good||BP Clonase Enzyme||Enzyme|
5X reaction buffer