plasmid DNA running on agarose gel to the opposite direction - (Jul/27/2006 )
Someting strange happend to me...I loaded 1 kbp Molecular weight markers and my plasmids from minipreps on 1% agarose gel and run it as always. The markers run and separate very nicely but the plasmid DNA migrates to the negative pole and obviously gets out of the gel.
Do you have any explanation for it???
If the markers ran correctly, and your "plasmid" ran backwards in the same gel, then your plasmid DNA is not DNA.
Which way did the tracking dye in your plasmid samples run?
Are you assuming that the sample DNA ran backwards because there was none visible on the gel after staining? The other explanation for such an observation is that there was no plasmid DNA in your samples to begin with.
Which way did the tracking dye in your plasmid samples run?
Are you assuming that the sample DNA ran backwards because there was none visible on the gel after staining? The other explanation for such an observation is that there was no plasmid DNA in your samples to begin with.
The first time I run it, everything was OK except that I could not see any DNA. The second time, I noticed that it is running out of the well because of the tracking dye (that was running both to the front and some of it out of the gel, so I inverted the current and I do have DNA which I can see by UV. But for some reason this DNA is positively charged...?????
what are you using as loading buffer?
did you use a bead prep on the DNA?
Yes, the BioRad miniprep kit
Glycerol/Bromophenol blue/Xylene cyanol FF
and it runs perfectly well. It is the same that I use to load the MW markers
The only thing I can think of is that you're DNA is bound to something positive (proteins? beads?) and they're dragging the DNA towards the negative pole. It's curious that your tracking dye ran both ways (?) -- could it be a pH problem?