Category Archives: Gel Electrophoresis

Table: Migration of bromophenol blue and xylene cyanol on agarose gel in TBE and TAE electrophoresis buffer

Position of bromophenol blue and xylene cyanol in agarose gel in relation to the position of double standard DNA fragment in TAE (1x) and TBE (0.5 x) electrophoresis buffer. For example in 0.5% agarose gel, Bromophenol blue migrates at approximately 750 bp long double standard DNA fragment in TBE buffer and at approximately 1150 bp long double standard DNA fragment in TAE buffer.
Agarose gel, %	Bromophenol blue in TBE buffer (0.5x)	Xylene cyanol FF) in TBE buffer (0.5x)	Bromophenol blue in TAE buffer (1x)	Xylene cyanol FF in TAE buffer (1x)
0.5	750	13000	1150	16700
0.6	540	8820	850	11600
0.7	410	6400	660	8500
0.8	320	4830	530	6500
0.9	260	3770	440	5140
1.0	220	3030	370	4160
1.2	160	2070	275	2890
1.5	110	1300	190	1840
2.0	65	710	120	1040

Reference:

https://assets.thermofisher.com/TFS-Assets/LSG/manuals/MAN0012614_Gen_Recommend_DNA_Electrophoresis_UG.pdf

Properties	TBE	TAE	Remarks
Buffering capacity	High	Low	TAE become exhausted during extended or repeated electrophoresis
Migration of (double stranded) DNA	Slow	Fast	TAE has better conductivity
Resolution	High resolution long DNA fragments	High resolution for Short DNA fragments	TBE supports agarose cross-linking better than TAE
Enzymatic modification of gel purified DNA	Poor	Good	Borate from TBE buffer is an inhibitor of many commonly used enzymes in molecular biology
Recovery of DNA from agarose gel	Poor	Good	Borate from TBE buffer interacts with DNA
Integrity of DNA	High	Low	Borate from TBE buffer inhibits many enzyme activity including DNA modifying enzymes
Cost	Costly	Cheaper	Highly conc. TAE (50X) require less volume to be transported.
Active buffer	Tris – Borate	Tris -Acetate
Working conc for DNA electrophoresis	1X	0.5X

Agarose gel can be used for the separation of nucleic acids (DNA and RNA) and high molecular weight proteins or protein complexes.
The process of using agarose gel to separate or fractionate biomolecules under the influence of electric field is called agarose gel electrophoresis.
For moderate and large sized DNA, agarose gel is the most commonly used medium in most molecular biology laboratories.
As the name suggest, the agarose gel is prepared from agarose.
Chemically, agarose is a polysaccharide, made up of alternating residues of 1,3-linked β-D-galactopyranose and 1,4-linked 3,6-anhydro-α- L-galactopyranose.