There are several factors that lead to poor results for PFG markers. Here is a list of recommendations for ensuring a sharp high quality banding pattern:
- Use Chef DR-II system with a hexagon grid 120 degrees (do not use field inversion, 180 degrees).
- Use fresh 0.5X TBE for the running buffer and the gel.
- Use high quality water (Milli Q) for making the buffer and the gel.
- Load the marker as solid plug (never melt the plug).
- Load a thin slice of the marker, about 1 gradation, on the syringe (a thick plug will overload the well with DNA and lead to smeared blurry pattern).
- Always run the recommended program for each specific marker. The programs are optimized for resolving bands in the size range corresponding to each marker. Interchanging programs with different markers will often result in a poor resolution.
- Store the marker at -20°C.