I think you could get.. But is that the quality of the DNA that matters more?!

you will get clones with the reduced amounts of dna.

You will lose dna during precipitation to concentrate it. As you have a small volume of dna and you want to make it more concentrated you could just put the dna tube opened on a pcr block and evaporate some of the water/buffer off the sample leaving a more concentrated solution. 60c should do it but it would be best not to allow the sample to dry completely.

If your institute has a vacuum concentrator then that would be even better for concentrating your dna while keeping it frozen but as your method uses a 50ul reaction volume there is no need to concentrate as 35ul sample and 5ul reaction buffer and probably 1ul of enzyme still leaves you adding water to make up the reaction volume to 50ul

Thanks for the answers.

Paul Rutland Won't I lose any DNA if I warm up my sample in the pcr block?

Marco Pedretti I got my DNA from gel extraction, and because my primers have contamination, I can't do pcr on it again.

I was planning to concentrate my DNA, and since the recommended amount of DNA in a 25ul digestion rxn is 500ng, decrease the total rxn to 25ul. Do you think it's a good idea?

What if I do another gel extraction, get around 1ug of the DNA in total and then concentrate my sample by heating it up to get a sufficient amount for the digestion reaction. Would that be a good idea?

Paul Weber just the water will evaporate off. There may be a small amount of dried dna on the wall of the tube but that will redissolve on vortexing but concentration is unneccessary anyway in this case. I think the answer to your situation is to either cut in 40ul total reaction volume or 50ul and at the next stage it is the amount of dna that matters so just adjust the volume of digested dna to suite your methodology. It is unlikely that you will be using all of your material for cloning and it is the ratio of insert dna to plasmid that is the most important in cloning so as long as you know how much you end up with you should get good cloning

Paul Rutland Thanks a lot. Just out of curiosity, if heating the DNA sample doesn't damage the DNA itself, is it OK to elute the DNA in say 100-200ul of water to get a better yield, every time that I do column purification, and then heat up the sample to get my desired volume?