Protocol Online logo
Top : New Forum Archives (2009-): : Molecular Cloning

Gibson/SLIC question - (Mar/05/2014 )

I'm a bit confused regarding Gibson assembly or SLIC. Using an exonuclease should result in regression of both 5' ends, right? Why is in all schemes only one of them degraded ( )? Isn't there a chance that the whole PCR product will be eventually degraded? Or are they using modified primers?






Sure, that makes sense. What I mean is that both ends of the vector and the PCR product should be chewed away, such as


5'--------------------------------3'        goes to                      5'-----------------3'        and not    5'-------------------------3'      as they show in the picture. 

3'--------------------------------5'                           3'------------------5'                                      3'-------------5'


Why is the left arm of the DNA "protected" in the pic? ie why is only one DNA strand degraded?