E-Gel Buffer Compatability

The E-Gel manual recommends low salt buffers:

Important: Samples containing ≥50 mM NaCl, 100 mM KCl, 10 mM acetate ions, or 10 mM EDTA (i.e. certain restriction enzyme and PCR buffers) will cause loss of resolution on E-Gel® agarose gels. To obtain the best results, dilute samples which contain high salt levels 2- to 20-fold.

NEB Buffer 4 contains very high acetate concentrations:

1X NEBuffer 4: 20 mM Tris-acetate 50 mM potassium acetate 10 mM Magnesium Acetate 1 mM Dithiothreitol pH 7.9 @ 25°C

Whereas buffer 2 does not:

1X NEBuffer 2: 10 mM Tris-HCl 50 mM NaCl 10 mM MgCl2 1 mM Dithiothreitol pH 7.9 @ 25°C

Ligation buffer is also acceptable:

1X T4 DNA Ligase Reaction Buffer: 50 mM Tris-HCl 10 mM MgCl2 1 mM ATP 10 mM Dithiothreitol pH 7.5 @ 25°C

The symptom is extreme band smearing of gel bands.