TAE buffer 2833999 223824419 2008-07-06T00:19:07Z Achian 495171 '''TAE buffer''' is a [[buffer solution]] used in agarose [[electrophoresis]], typically for the separation of [[nucleic acids]] such as [[DNA]] and [[RNA]].<ref>Ogden, R.C., and Adams, D.A., Electrophoresis in agarose and acrylamide gels. Methods Enzymol., 152, 61-87 (1987).</ref> It is made up of [[Tris-acetate]] buffer, usually at pH 8.0, and [[EDTA]], which sequesters divalent cations. TAE has a lower buffer capacity than [[TBE buffer|TBE]] and easily can become exhausted, but linear, double stranded DNA runs faster in TAE. ==Uses== TAE buffer is used as both a running buffer and in agarose gel.<ref>Sambrook, Fritsch, and Maniatis (1989) ''Molecular Cloning: A Laboratory Manual'', 2nd ed., Cold Spring Harbor Laboratory Press, Cold Spring Harbor, New York, volume 3, apendices B.11 and B.23</ref> Its use in denaturing gradient [[gel electrophoresis]] methods for broad-range mutation analysis has also been described.<ref>Hayes, V.M. et al., Improvements in gel composition and electrophoretic conditions for broad-range mutation analysis by denaturing gradient gel electrophoresis. Nucleic Acids Res., 27(20), e29 (1999).</ref> TAE has been used at various concentrations to study the mobility of DNA in solution with and without [[sodium chloride]].<ref>Stellwagen, E., and Stellwagen, N.C., The free solution mobility of DNA in Tris-acetate-EDTA buffers of different concentrations, with and without added NaCl. Electrophoresis, 23(12), 1935-1941 (2002).</ref> However, high concentration of sodium chloride (and many other salts) in a DNA sample retards its mobility. This may lead to incorrect interpretations of the resulting DNA banding pattern. Compared with [[TBE buffer]], TAE buffer offers advantages in subsequent enzymatic applications for the DNA sample. For example, if a DNA sample is going to be used in a cloning experiment, the step that follows its running on a agarose gel is to ligate (covalently link) to a cloning vector (most likely a plasmid). DNA sample from TAE buffer is suitable for this purpose, while DNA from TBE buffer is not. Borate in the TBE buffer is a strong inhibitor for many enzymes. By the way, this (enzyme inhibiting) property made [[TBE buffer]] very popular in its realm for two reasons. First, DNA samples run in a TBE buffer can better keep its integrity. The other main reason is that the purpose for many agarose gel electrophoreses is to analyze the size of DNA molecules. In particular, the kinds of DNA analyses shown on TV and other public media are more likely run in TBE buffer. ==Preparation== For 1 litre of 50x TAE buffer use: - 242 g Tris base (2-amino-2-hydroxymethyl-propane-1,3-diol) (= 2 mole) - 57.1 ml glacial acetic acid (= 100% acetic acid) (57.19 ml = 1 mole) - 100 ml 0.5 M Na<sub>2</sub> EDTA (pH 8.0) - H<sub>2</sub>O up to 1000 ml To prepare 0.5 M Na<sub>2</sub> EDTA (pH 8.0) add 186.1 g of disodium ethylenediaminetetraacetate x 2H<sub>2</sub>O to 800 ml of H<sub>2</sub>O. Stir vigorously. Adjust the pH to 8.0 with NaOH (ca. 20 g of NaOH). Sterilize by autoclaving. Hint: The disodium salt of EDTA will not go into solution until the pH of the solution is adjusted to ca. 8.0 by the addition of NaOH. ==Reference== <references/> ==See also== * [[TBE buffer]] [[Category:Buffers]] {{biochem-stub}} [[de:TAE-Puffer]] [[fr:Tris, Acétate, EDTA]] [[pt:Tampão TAE]]