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Gibson Assembly Cloning issues

Gibson cloning assembly primers

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#1 Natalia KM

Natalia KM


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Posted 10 September 2021 - 10:28 PM

Hi there,


Can someone please help me with Gibson Assembly of my ~4KB insert into a pAAV vector.


I have followed all the steps but having problems.


Help with primer design, reaction conditions etc and if anyone could look at my specific sequences would be really helpful. I can send sequences etc if anyone is willing to look.


Prior to the PCR reactions, do the vectors containing the insert or the destination vector need to be linearised? I did not think that was necessary so did not do that.


Any help would be greatly appreciated. 



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