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the need for the donor vector as intermediate step

gateway entry clone

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#1 daisato

daisato

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Posted 23 March 2012 - 09:17 AM

Hi all,
I'm just starting to get myself acquanted with the Gateway cloning system. One question that came to my mind while studying this system was:
If you create your destination vector with attP sites (rather than attR sites), can you then clone your attB flanked PCR product directly into your destination vector, eliminating the necessity for the donor vector and the extra LR clonase step? The need for the two recombination steps to reach the final product isn't immediately clear to me. I know that by doing so, you create the entry clone that can be swapped into any destination vector, but does this gain you really that much profit/time winning/less laborious that just clone the PCR product to the destination vector directly? Especially if there will probably not be that much need to be able to quickly/easily clone the product to multiple different destination vectors.

I'm not sure whether this is a completely ignorant question, but it just crossed my mind. I hope someone can explain the benefits of this to me. Thanks a lot in advance!





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