10. When you make your agarose gel, you will need to make 50 ml of 1% (w/v) agar
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Question
10. When you make your agarose gel, you will need to make 50 ml of 1% (w/v) agarose in 1X TAE. How much agarose will you require? [you will make this later, not on Solution day] When adding your DNA samples to the agarose gel, you will need to add loading dye to the samples. Loading dye contains: • glycerol, which helps the samples sink into the sample wells of the gel; and • tracking dyes so that you have a way of estimating how far the samples have travelled through the gel during electrophoresis. The recipe for 6X loading dye is as follows: • 0.25% bromophenol blue (w/v) • 0.25% xylene cyanol (w/v) • 30% glycerol in water (v/v)
11. Provide a recipe for 10 ml of 10X loading dye, using the information above. (Important tip: Remember that you have already made 30ml of a 50% stock solution of glycerol. I recommend determining the concentration of glycerol required for 10X loading dye, then figuring out how much of your 50% glycerol stock will be needed to make 10ml of 10X loading dye.)
Solution to make
Exact pH of final solution (if appropriate)
Final container for solution
Completed (Y or N)
100 ml of 0.9% (w/v) NaCl
100 ml of 1.0M Tris, pH 7.6
40 ml of 50X TAE
Solution to make
Exact pH of final solution (if appropriate)
Final container for solution
Completed (Y or N)
100 ml of 0.9% (w/v) NaCl
100 ml of 1.0M Tris, pH 7.6
40 ml of 50X TAE
Explanation / Answer
10. The electrophoresis is the movement of charged particles under the electric field influence. The agarose gel is the stabilizing medium used for electrophoretic separation of nucleic acids. The agarose gel has resolving power which depends on pore size which is dependent on the concentration of agarose dissolved. So the agarose concentration is very crucial.
So to prepare 50 ml 1% (w/v) agarose in 1X TAE we require 0.5 g of agarose in 50 ml 1XTAE.
The recipe for 6X DNA loading dye (10 ml)
- 25 mg bromophenol blue to 6.7 ml of ddH2O
- 25 mg of xylene cyanol FF
- 3.3 ml of glycerol
This is used to prepare tracking dye.
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