What was the 260/280 ratio on the nanodrop? Personally I much prefer fluorometric quantitation, as you know that you are only detecting dsDNA and not other residual proteins, etc, remaining from the extraction. You are using a fairly crude extraction method too, 20 min at 100 degrees is certainly going to denature any naked dsDNA, which may also explain what you are seeing on the gel, especially if you are using dsDNA dyes like ethidium bromide.

Nanodrop is a spectrophotometer, it does not quantify crude DNA extract correctly.

Thank you for your opinions. Mr. Payne, look at the 260/280 ratios and their corresponding DNA concentrations (or whatever they may be). I ran the sample again today on a 0.8% agarose gel, same running conditions. The marker turned out very well but for my samples none even appeared in the wells.

What sort of dye are you staining your gel with?

Ecodye Nucleic acid staining solution from Biofact (I think this is a Korean product or Japanese product). The marker stains absolutely well

Personnally, I don't think that skimmed milk vs glycerol is the problem since you cultured the isolate on BHI plate before performing DNA extraction. Do you have the possibility to try another DNA extraction protocol? Personally, I use Qiagen kit and it works very well. Also, could you post your gel picture?

Good luck,

Simon