DNA shows on Nanodrop but not on electrophoresis - (Nov/05/2014 )

Hello all,

I am extracting (or trying to) DNA from Ascaris and something odd is happening. For the extraction i use a lysis buffer with SDS, proteinase K followed by phenol:chloroform, isopropanol extraction. Pretty standard and i have tried several variations and also the Qiagen DNeasy blood and tissue kit.

Problem is: when i quantify the samples on Nanodrop it shows i have DNA (about 100ng/uL) and with good quality. When i run an agarose gel (0.7%; 0.5ug/mL etbr) i don't see any bands on Ascaris samples. I run the extractions in parallel with human DNA and those works perfectly.

Have anyone seen anything like this? Any clues?

Thanks a lot.

Nanodrop and similar spec methods don't tell you anything about the DNA quality, just whether you are likely to have extracted DNA (RNA absorbs pretty much in the same spectrum) and how much.

How much DNA are you running on the gel -not enough and you won't see it?

It could be a number of things, including that the DNA is getting stuck in the wells of the gel.

It could be that the DNA is heavily degraded (the gut is full of DNases), stain a gel after running (rather than adding EtBr to the gel) and check for smear, especially in places where you might see ethidium shadow.

It could be that what the nanodrop is seeing isn't DNA but something else (perhaps another sugar based compound such as mucus if the gut contents aren't fully washed off).

Hello bob1 thanks for the reply. 

By DNA quality i meant the ratio A260nm/A280nm and A260nm/A230nm.

The DNA is not getting stuck on the wells. 

I am not sure that the reading on Nanodrop is something other than nucleic acid because the results with the Qiagen kit (which as you probably know binds DNA on silica) are the same. 

I´ll try staining the gel after the run. Hopefully it will give some clues.

Again, thanks.