Preparation of 6X DNA Loading Dye (Bromophenol blue, xylene cyanol FF and Sucrose) Alternative Method

OVERVIEW

This method uses 40% sucrose solution instead of sucrose powder. This method is convenient and easy, and does not involve adjustment of volume.

REQUIREMENTS

Reagents and solutions
Bromophenol blue
Xylene cyanol FF
40% (w/v) Sucrose solution (sterile)
Deionized / Milli-Q water

Equipment and disposables
15-ml screw-cap graduated polypropylene centrifuge tube
Centrifuge (for 15 ml tube) 
Tube rotator (optional) 
Vortex mixer (optional)

COMPOSITION

Composition of 6X DNA loading dye 
0.25% (w/v) bromophenol blue
0.25% (w/v) xylene cyanol FF
40% (w/v) sucrose

Composition of 1X DNA loading dye 
0.042% (w/v) bromophenol blue 
0.042% (w/v) xylene cyanol FF
6.67% (w/v) sucrose

OBJECTIVE
Preparation of 10 ml of 6X DNA loading dye containing bromophenol blue, xylene cyanol FF and Sucrose

PREPARATION

To prepare 10 ml of 6X DNA loading dye, weigh out 25 mg bromophenol blue and 25 mg xylene cyanol FF. Transfer it to a 15-mL screw-capped graduated tube. Add 10 ml of 40% sucrose solution. Mix vigorously using a vortex mixer or tube rotator to dissolve bromophenol blue and xylene cyanol FF in 40% sucrose solution. 

Note
This method is simple and convenient as it does not involve readjustment of solution volume. There will not be any significant change in the volume of the 40% sucrose solution when you dissolve small amounts of  bromophenol blue and xylene cyanol FF in it.

Tip
We recommend using disposable nuclease nuclease-free (DNase- and RNase-free)15 ml screw-capped graduated tubes instead of beaker as all the procedure can be done in a single tube. 

STORAGE
Store the solution in the freezer (-20°C). Solution can be stored at 2 – 8°C for a few weeks.

Precautions
Sucrose-containing loading dyes are prone to mold-like growth. Avoid keeping it in the refrigerator or room temperature. 

Tips
It is recommended to store the solutions in small aliquots (1 ml) at -20°C.

APPLICATIONS
This solution is used for loading DNA samples onto non-denaturing gels.

Follow the table to prepare 6X agarose gel loading dye of various volumes.
Reagents / Volume5 ml10 ml25 ml50 ml100 ml
Bromophenol blue12.5 mg25 mg62.5 mg125 mg250 mg
Xylene cyanol FF12.5 mg25 mg62.5 mg125 mg250 mg
40% Sucrose solution5 ml10 ml25 ml50 ml100 ml

CALCULATOR

Use Calculator to calculate the amount of different components of 6X DNA loading dye

Volume of the 6X DNA loading dye: ml
ml(Change the volume of the solution)

Amount of Bromophenol Blue: mg

Amount of Xylene Cyanol FF: mg

Amount of 40% Sucrose: ml

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