Problem in Ligation - (Jun/28/2010 )

I agree. I too believe that the 1:1 mol ratio of is best. Equal number of molecules.

What do you mean with 1:1 mol ratio?

You mean you need to have the same amount of mols of the vector and the insert?

And how do you calculate how much mole of each you have?
Normally you only know how much bases it is?

josse on Sat Jul 31 15:36:17 2010 said:

Yes! 1:1 mol ratio means same number of moles (and hence molecules) of insert and vector and it is preferred because 1 molecule of insert ligates to 1 molecule of vector. You need not know absolute number of moles of each, rather you are interested only in their ratio and while doing so it has to be assumed that the molecular weight of a fixed length of DNA having any composition would be the same (same GC content).
If length (number of bases) of insert is 'x' and that of vector is '3x' (3 times larger than insert) than a particular weight (for example, in nanograms) of insert will have 3 times the number of moles in the same weight of vector. Although ready-made calculators are available at websites like this, I will recommend to get your moles, molar and molecular biological concepts cleared if it is still difficult for you.
Good luck!