TBE or Tris/Borate/EDTA, is a
buffer solution
A buffer solution is a solution where the pH does not change significantly on dilution or if an acid or base is added at constant temperature. Its pH changes very little when a small amount of strong acid or base is added to it. Buffer solution ...
containing a mixture of
Tris base,
boric acid
Boric acid, more specifically orthoboric acid, is a compound of boron, oxygen, and hydrogen with formula . It may also be called hydrogen orthoborate, trihydroxidoboron or boracic acid. It is usually encountered as colorless crystals or a white ...
and
EDTA
Ethylenediaminetetraacetic acid (EDTA), also called EDTA acid, is an aminopolycarboxylic acid with the formula . This white, slightly water-soluble solid is widely used to bind to iron (Fe2+/Fe3+) and calcium ions (Ca2+), forming water-solubl ...
.
In molecular biology, TBE and
TAE buffers are often used in procedures involving
nucleic acids
Nucleic acids are large biomolecules that are crucial in all cells and viruses. They are composed of nucleotides, which are the monomer components: a 5-carbon sugar, a phosphate group and a nitrogenous base. The two main classes of nucleic a ...
, the most common being
electrophoresis
Electrophoresis is the motion of charged dispersed particles or dissolved charged molecules relative to a fluid under the influence of a spatially uniform electric field. As a rule, these are zwitterions with a positive or negative net ch ...
.
Tris
Tris, or tris(hydroxymethyl)aminomethane, or known during medical use as tromethamine or THAM, is an organic compound with the formula (HOCH2)3CNH2. It is extensively used in biochemistry and molecular biology as a component of buffer solutions ...
-acid solutions are effective buffers for slightly basic conditions, which keep DNA deprotonated and soluble in water. EDTA is a
chelator of divalent
cations
An ion () is an atom or molecule with a net electrical charge. The charge of an electron is considered to be negative by convention and this charge is equal and opposite to the charge of a proton, which is considered to be positive by convent ...
, particularly of
magnesium
Magnesium is a chemical element; it has Symbol (chemistry), symbol Mg and atomic number 12. It is a shiny gray metal having a low density, low melting point and high chemical reactivity. Like the other alkaline earth metals (group 2 ...
(Mg
2+). As these ions are necessary co-factors for many enzymes, including contaminant
nucleases
In biochemistry, a nuclease (also archaically known as nucleodepolymerase or polynucleotidase) is an enzyme capable of cleaving the phosphodiester bonds that link nucleotides together to form nucleic acids. Nucleases variously affect single and ...
, the role of the EDTA is to protect the nucleic acids against enzymatic degradation. But since Mg
2+ is also a co-factor for many useful DNA-modifying enzymes such as
restriction enzyme
A restriction enzyme, restriction endonuclease, REase, ENase or'' restrictase '' is an enzyme that cleaves DNA into fragments at or near specific recognition sites within molecules known as restriction sites. Restriction enzymes are one class o ...
s and
DNA polymerase
A DNA polymerase is a member of a family of enzymes that catalyze the synthesis of DNA molecules from nucleoside triphosphates, the molecular precursors of DNA. These enzymes are essential for DNA replication and usually work in groups to create t ...
s, its concentration in TBE or TAE buffers is generally kept low (typically at around 1
mM).
According to studies by Brody and Kern, sodium boric acid is a superior and cheaper conductive media for most DNA
gel electrophoresis
Gel electrophoresis is an electrophoresis method for separation and analysis of biomacromolecules (DNA, RNA, proteins, etc.) and their fragments, based on their size and charge through a gel. It is used in clinical chemistry to separate ...
applications.
Recipe (1 liter of 5X stock solution)
*54 g of
Tris base (CAS# 77-86-1, free base)
*27.5 g of
boric acid
Boric acid, more specifically orthoboric acid, is a compound of boron, oxygen, and hydrogen with formula . It may also be called hydrogen orthoborate, trihydroxidoboron or boracic acid. It is usually encountered as colorless crystals or a white ...
(CAS# 10043-35-3)
*20 ml of 0.5 M
EDTA
Ethylenediaminetetraacetic acid (EDTA), also called EDTA acid, is an aminopolycarboxylic acid with the formula . This white, slightly water-soluble solid is widely used to bind to iron (Fe2+/Fe3+) and calcium ions (Ca2+), forming water-solubl ...
(CAS# 60-00-4) (
pH 8.0)
Adjust pH to 8.3 by HCl.
TBE can be diluted to 1X prior to use in electrophoresis, 0.5x is acceptable as well. Higher concentrations will result in poor results due to excessive heat generation.
See also
*
LB buffer
LB buffer, also known as lithium borate buffer, is a buffer solution used in agarose electrophoresis, typically for the separation of nucleic acids such as DNA and RNA. It is made up of Lithium borate (lithium hydroxide monohydrate and boric acid) ...
, lithium borate buffer, a similar buffer containing lithium ions in place of Tris
*
TAE buffer
TAE buffer is a buffer solution containing a mixture of Tris base, acetic acid and EDTA.
In molecular biology, it is used in agarose electrophoresis typically for the separation of nucleic acids such as DNA and RNA. It is made up of Tris-acetate ...
, a similar buffer containing acetic acid in place of boric acid
Notes
References
{{DEFAULTSORT:Tbe Buffer
Buffer solutions