I used this method, and works great. The most important thing is the ratio of DN A amount has to be correct, Use less DNA than regular cloning method. If you failed, I would say most likely you used too much DNA. I would suggest when use PCR product or oligos, try 1:10, 1:100, 1:1000 dilutions, one of them will work


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> On Feb 18, 2019, at 16:43, Hanke van der Wel <[log in to unmask]> wrote:
> 
> I tried using the LIC method to ligate 3 fragments into a KpnI-linearized vector using the HiFi DNA Assembly Master Mix from NEB, but was unsuccesful. 
> No clones on the plate after transfection, and no ligated fragments detected with pcr.
> 
> Has anyone tried this method with Dicty DNA (or maybe another form of Gibson assembly), and if so, do you have hints/suggestions?
> 
> Thanks!
> 
> 
> 
> Hanke van der Wel
> Research Professional IV
> Biochemistry & Molecular Biology
> University of Georgia
> 120 E. Green Street
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> Athens, GA 30602
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