Tris-acetate buffer

Agarose gel electrophoresis is a conventional method for separating and identifying nucleic acids in molecular biology laboratories. Nucleic acids are amphoteric electrolytes with an isoelectric point of pH 2-2.5. In conventional electrophoresis buffers (pH about 8.5), nucleic acid molecules are negatively charged. The charge moves towards the positive pole in the electric field. Through electrophoresis technology, RNA molecules of different sizes and conformations can be separated, and by observing the number, size and shape of the bands, the integrity of the RNA molecules can be judged. In this process, the main function of the electrophoresis buffer is to maintain the pH and make the solution have a certain conductivity. TAE (Tris-acetic acid buffer) and TBE (TRIS-boric acid electrophoresis buffer) are commonly used buffers. What is the difference between them? TAE (Tris-acetate buffer) Advantage: ① For DNA fragments larger than 13kb, TAE buffer can achieve better separation results. ② In TAE buffer, the supercoiled state of DNA can better maintain its true relative molecular mass during electrophoresis. ③Suitable for recovering DNA fragments and subsequent enzyme digestion reactions, etc. shortcoming:   The buffer capacity is small, and TAE buffer may gradually lose its pH buffering capacity during long-term electrophoresis, resulting in changes in pH value. TBE (TRIS-boric acid electrophoresis buffer) Advantage: ① The buffering capacity is relatively stronger, the pH value can be kept constant even during long-term electrophoresis, and it is not easy to cause overheating in the electrophoresis tank. ② Suitable for electrophoretic separation of smaller fragments, such as fragments less than 1kb. Its high resolution makes it easier to distinguish molecules of different sizes, thereby increasing the accuracy of experiments. shortcoming: The boric acid component contained may affect the recovery efficiency of RNA and DNA and subsequent enzymatic reaction experiments.