I'm having an (ongoing) issue with possibly my DNA extractions and gel electophoresis. Some information: I am extracting DNA from animal tissue (mouse tail clip) in order to genotype our mouse colony. More often than not when I run a gel the bands of at least half of my samples are very faint or completely absent. Recently (this past week or so) absolutely none of my samples can be visualized on the gel. I am not sure what the problem is or what I am (have been) doing wrong. I thought it was a problem with my DNA extractions because I noticed that with almost all of my samples the DNA pellet never seems to dissolve into the TE buffer. Once re-suspended I heat and shake my samples overnight at 37C, but the pellet never seems to dissolve. I tried adding more TE buffer to each of my samples but this did not seem to help.
To determine whether it was my samples or my primers I ran a PCR using the same samples but different primers then ran those samples on a gel. When I did this the bands came out as we expected them to. This made me think that maybe the problem is not with the extractions (in spite of the fact that the DNA never seems to dissolve into the TE buffer) and that I may be doing something wrong when preparing the PCR.
Both the master mix and primers are new so I now think I may not be using the right amount of DNA and/or primer for the PCR rxt. As you can imagine this is very frustrating and being that I am relatively new to all this, any advice would be super helpful...
some additional information just in case: all of the buffers I have been using (i.e., lysis buffer for tail digestion and TE for re-suspension) were made within the past month, so they are also relatively new. We run our samples on a 1.5% agarose gel stained with ethidium bromide at around 45 mV.