May 27, 2022

Public workspacePreparation of 2% Agarose Gel for Electrophoresis

This protocol is a draft, published without a DOI.
  • 1Mboalab, Beneficial Bio
Nadine Mowoh
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Protocol Citation: Stephane Fadanka, Mujar Minette Shalo, Nadine Mowoh 2022. Preparation of 2% Agarose Gel for Electrophoresis. protocols.io https://protocols.io/view/preparation-of-2-agarose-gel-for-electrophoresis-b87rrzm6
License: This is an open access protocol distributed under the terms of the Creative Commons Attribution License,  which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited
Protocol status: Working
We use this protocol and it’s working
Created: May 11, 2022
Last Modified: May 27, 2022
Protocol Integer ID: 62417
Abstract
The agarose gel consists of microscopic pores that act as a molecular sieve which separates molecules based upon the charge, size and shape.Agarose gel electrophoresis can also be used to separate other charged biomolecules such as RNA and proteins.Agarose is isolated from the seaweed genera Gelidium and Gracilaria and consists of repeated agarobiose (L- and D-galactose) subunits.The concentration of agarose in a gel depends on the sizes of the DNA fragments to be separated, with most gels ranging between 0.5%-2%. 
Materials
Reagents

Agarose powder(electrophoresis grade)
1xTBE
Nucleic acid Stain(SafeView)

Materials and Equipment

Weighing boat
Electronic balance
Spatula
Electrophoretic gel tank and components(BlueGel)
Microwave
Beaker 
Micropipette P-10
10ul micropipette

Safety warnings
The entire process is generally safe but it is advisable to wear proper protective clothing before staring.
Measuring Agarose powder
Measuring Agarose powder
22m
22m
Accurately weigh Amount0.5 g of Agarose- electrophoresis grade (Cas 9012-36-6) into a weighing boat using a clean spatula and transfer the powder into a 50ml or 100ml beaker.

Measure Amount25 mL of 1xTBE (diluted from a 10x stock as described in the citation below )and dispense into the beaker
CITATION
Nadine Mowoh, Jenny Molloy. Preparing 10x TBE Electrophoresis buffer.
LINK
dx.doi.org/10.17504/protocols.io.j8nlkkok5l5r/v1


Mix by swirling gently and transfer the beaker into the microwave at medium high to heat and boil for Duration00:01:00

1m
Turn off the microwave and take out the beaker , allow it to cool for Duration00:01:00 atTemperatureRoom temperature , just enough that you can conveniently hold the beaker in your palm.

1m
Pipette Amount2.5 µL of nucleic acid gel stain (10000x) into the gel while it's still liquid and swirl gently to mix
Note
Avoid swirling too hard as it might generate bubbles that will make your gel rough and affect migration of nucleic acid


Assemble the electrophoretic gel casting tray on a level surface and put the combs in place
Pour the gel gently into the tray to avoid bubbles
Leave gel to set for Duration00:20:00 to polymerise

20m
Carefully remove the comb from the solidified gel to have the sample wells
Transfer the gel tray with the solidified gel to a horizontal electrophoretic gel tank and add about Amount35 mL of TBE buffer (depending on the size of your gel tank) to completely cover the gel. Load the nucleic acid samples into the wells and allow them run.

Citations
Step 2
Nadine Mowoh, Jenny Molloy. Preparing 10x TBE Electrophoresis buffer
dx.doi.org/10.17504/protocols.io.j8nlkkok5l5r/v1