AMRESCO AGAROSES - Code 0710, 0815, X174, E434, E776, J234

Directions for Use

AMRESCO has received several inquiries regarding the proper preparation of our Agaroses for electrophoretic purposes. Proper heating of your agarose/buffer mixture is important in obtaining a gel mixture that is easy to cast and that will polymerize correctly. We would like to take this opportunity to provide users with some helpful hints in preparing Agarose I or any of AMRESCO’s other Ultra Pure agaroses.

In general, heating times are shorter for AMRESCO's Agaroses than other brands of agarose.
When agarose is placed in a buffer such as TAE (Tris/Acetate/EDTA) or TBE (Tris/Borate/EDTA), it is generally insoluble. However, when this agarose solution is heated, the agarose particles become hydrated and thus go into solution. This hydration process is time-dependent, and different types of agarose will have varying hydration points. AMRESCO’s Agaroses are extremely pure and comprised of ultra-fine particles.

 This ultra-fine structure is important in providing a porous, highly resolving matrix for electrophoretic applications. As a result of this structure, AMRESCO Agaroses will have a faster rate of hydration than many other types of agaroses. End-users who have used other brands of agarose in the past may mistakenly boil AMRESCO Agarose much longer than is needed, which results in a thick gelatinous solution that is difficult to cast and brittle when polymerized.

Helpful Hints For Preparing AMRESCO’s Agaroses

Preparation of a typical 1% agarose gel 1X TBE buffer.

  • In an appropriate container (an Erlenmeyer flask at 2-4X the volume of the desired gel volume is optimal), slowly add agarose crystals to your buffer solution while gently swirling. This will help to eliminate clumping of the agarose. Record the weight of the flask containing the buffer and agarose.

  • Heat the solution in a microwave on high power for 30 seconds. (For smaller or larger volumes, increase or decrease heating times proportionally to volume size). Heating times will vary depending on your microwave oven (wattage), size of the flask used and the % agarose.

  • Swirl the agarose solution gently to re-suspend the particles.

  • Heat the solution another 30 seconds on high power, remove and swirl the agarose solution.

  • Place the solution back in the microwave and heat on high power until the solution just starts to boil (boiling point will probably take 10-35 seconds). Use caution when handling the hot flask. Microwaved solutions may become superheated and can boil vigorously when moved or touched. After removing the boiling solution from the microwave oven, allow to cool briefly (1-2 minutes) at room temperature, then gently swirl the solution to release entrapped air (some air bubbles will remain).

  • Place the agarose solution back in the microwave, heat on high power and let the solution boil for approximately 15 seconds. Inspect the solution for agarose crystals (they will appear as floating ÒlensesÓ) while gently swirling. If there are particles present, repeat this step until all crystals are dissolved.

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  • Once the agarose is completely in solution, again weigh the flask to check for water loss by evaporation. Replenish with water as necessary (until the weight of the flask and its contents equal the original weight). Gently swirl the solution.

  • In general, it is advisable to allow any agarose solution to cool to ( 50-55(C on the lab bench prior to pouring into a prepared apparatus. This is conductive to a more uniform pore size and will prevent the warping of your gel apparatus. Before pouring the gel, gently swirl the agarose solution to help dissipate most of the remaining air bubbles.

  • Pour the gel into the prepared casting unit. Usually, horizontal gels should be 3-5mm thick. Immediately after pouring, check to see that there are no air bubbles under or between the teeth of the gel comb.

  • Allow the gel to completely polymerize at room temperature (about 30-45 minutes) before running your samples. For further information on agarose gel electrophoresis, see Sambrook et al. (Chapter 6).

Once the boiling point has been reached, observe your solution after each 10-15 second boiling interval very closely (depending on concentration and volume).

  • Gently swirl your agarose solution at least twice before you bring the solution to a boil.

  • Once the boiling point has been reached, observe your solution after each 10-15 second boiling interval very closely (depending on concentration and volume).

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Product Gel % Optimal Separation (bp) Recommended Buffer

Agarose Iª

(analytical)

 0.80 800-22,000 TAE
1.00  500-10,000 TAE/TBE
1.20  400-7,000 TAE/TBE
2.00  250-5,000 TBE
Agarose IIª

(preparative or

analytical) (compatible

with in-gel manipulation)

 0.50  2,000-20,000 TAE*/TBE
0.80 500-15,000 TAE*/TBE
1.00  200-10,000 TAE*/TBE
 2.00  100-1,000 TAE*/TBE

Agarose IIIª

(analytical)

0.50  10,000-40,000 TAE
0.70 1,000-20,000  TAE
Agarose IVª

(analytical)

 0.50  10,000-2,000,000  TAE
 1.00  3,000-500,000   TAE
1.20  1,000-250,000   TAE

Agarose 3:1ª

HRB

(analytical)

1.00  500-2,000  TBE
2.00 250-750  TBE

3.00

 125-500 TBE

4.00 

20-250 TBE
Agarose SFRª

(preparative or 

analytical) (compatible 

with in-gel manipulation)

 2.00   500-2,000 TAE*/TBE
3.00   100-1,000 TAE*/TBE
4.00   50-500 TAE*/TBE
5.00   20-250 TAE*/TBE

 

Tableau de recommandation des agaroses Amresco en fonction de vos applications :

Resolve DNA fragments (50 – 2,000 bp)

Agarose 3:1 HRB and Agarose SFR

Resolve DNA fragments (1.0 – 22 kb)

Agarose I

Resolve DNA fragments (1.0 – 25 kb)

Agarose II

Resolve DNA fragments (>10 kb)

Agaroses III and IV

Southern and Northern Analysis

Agaroses I, III, IV and 3:1 HRB

Pulsed-field Gel Electrophoresis (PFGE)

Agaroses III and IV

Polymerase Chain Reaction* (PCR), in-gel

Agarose II and Agarose SFR

Cloning, in-gel

Agarose II and Agarose SFR

Restriction Digestion, in-gel

Agarose II and Agarose SFR

Sequencing, in-gel

Agarose II and Agarose SFR

DNA labeling, in-gel

Agarose II and Agarose SFR

 

 

 

 

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Vos contacts:

Etienne Boireau - Olivier Cadas - Olivier Alméras

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