band distortion in DNA gel electrophoresis - (Oct/30/2008 )

Hi all,

I am working with ISSR markers, which produce polymorphic DNA bands on the gel (the method is technically similar as RAPD). Those ´banding patterns´ sometimes show some uneven migration between samples. In attached picture of 2 different ISSR primers you can see samples SC 8 and SC 9 having nearly the same banding pattern (samples of closely related plants), but SC 9 is shifted downwards. The shift is also visible in case of a band, which I consider to be homologous among samples - marked by green line.

Another problem are U-shaped bands (see gel on the right side). Those bands appear especially when higher annealing temperature in PCR is used (increasing annealing temperature is main tool for ISSR-PCR optimization when banding patterns are poorly resolved). I use 1.8% agarose gels, run at 80V / 5hrs in 1x TBE, samples are stained by loading buffer+SybrGeen.
Please, does anybody have some idea?

Thanks!

Jirka