Concentration of DNA in Ligations???

M C Behrendt M.C.Behrendt at shef.ac.uk
Mon Jul 22 08:18:51 EST 1996


Does anyone know what the optimum concentration of DNA in a ligation 
reaction is (NOT the ratio of vector to insert), and what the ideal volume
per eppendorf should be.  At the moment I'm using approx 3-4ug of DNA (insert, 
660bp plus vector, 3674bp, 2:1 ratio) and 1-2 units of Ligase (+ buffer, from 
Gibco) in a total volume of 100ul which is aliquoted out in 10ul amounts for 
an overnight incubation at 11 or 16C.  My problem is that I need this to work 
with a very high efficiency in order to construct a library - but - it's not 
working.

Any suggestions would be appreciated

Mark





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