Bio-Rad DCode™ Universal Mutation Detection System - Manual (Page 19)

For denaturing solutions less than 100%, use the volumes for acrylamide, TAE and water
described above in the 100% Denaturing Solution. Use the amounts indicated below for urea
and formamide.
Denaturing Solution 10% 20% 30% 40% 50% 60% 70% 80% 90%
Formamide (ml) 4 8 12 16 20 24 28 32 36
Urea (g) 4.2 8.4 12.6 16.8 21 25.2 29.4 33.6 37.8
10% Ammonium Persulfate
Reagent
Amount
Ammonium persulfate 0.1 g
dH
2
O 1.0 ml
Store at –20°C for about a week.
DCode Dye Solution
Reagent
Amount Final
Concentration
Bromophenol blue
0.05 g 0.5%
Xylene cyanol 0.05 g 0.5%
1x TAE buffer 10.0 ml 1x
Store at room temperature. This reagent is supplied in the DCode electrophoresis reagent
kit for DGGE, CDGE.
2x Gel Loading Dye
Reagent
Amount Final
Concentration
2% Bromophenol blue
0.25 ml 0.05%
2% Xylene cyanol 0.25 ml
0.05%
100% Glycerol 7.0 ml
70%
dH
2
O 2.5 ml
Total volume 10.0 ml
Store at room temperature.
1x TAE Running Buffer
Reagent
Amount
50x TAE buffer 140 ml
dH
2
O 6,860 ml
Total volume 7,000 ml
Gel Volumes
Linear Denaturing Gradient Gels
The table below provides the required gradient delivery system setting per gel size desired.
The volume per syringe is for the amount required for each low and high density syringe, and
the volume adjustment setting sets the gradient delivery system for the proper delivery of
solutions. The 7.5 x 10 cm and 16 x 16 cm size gels are recommended for the perpendicular gel
formats, whereas the 16 x 10 cm and 16 x 16 cm gel formats are recommended for parallel
denaturing gels. The volume per syringe requires a larger volume of reagent than the volume
setting indicates, because the excess volume in the syringe is needed to push the correct amount
of gel solution into the gel sandwich.
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