分生实验（Meristematic experiment）.doc

分生实验（Meristematic experiment） 1. What is the role of the various reagents used in this experiment? Answer: NE solution: metal ion chelating agent, inhibit the activity of DNase; Protease K: broad-spectrum protease, which can maintain high activity in the presence of SDS and EDTA, degrade protein, and dissociate DNA; SDS: a. Anionic type remover, dissolving membrane and lipid composition of nuclear membrane; B. denaturation of protein; C. Destruction of membrane and nuclear membrane; Inhibition of nucleic acid enzymes; TE saturated phenol: remove impurities such as protein. Phenol has a strong denaturation effect on the protein, which can change the protein in the sample and remove it by centrifugation. In order to prevent its influence on subsequent experiment, the residual phenol was removed by chloroform extraction. Chloroform: a. Chloroform is not as modified as phenol, but it can be used to enhance the effect of protein. B. chloroform can accelerate the separation of organic and water phases, remove plant pigment and sucrose, and inhibit nucleic acid enzyme activity; C. Removal of residual traces of phenol in DNA aqueous solution; Isoamyl alcohol: reduce the production of bubbles during operation; Ethanol: in the presence of monovalent cation, ethanol can precipitate DNA. Sodium acetate: provide univalent cation; EDTA: inhibition of DNA enzyme activity; 2. What are the precautions during DNA extraction? A: 1) the TE saturated reagent, chloroform/isopentyl alcohol used in this experiment is an organic reagent that is more important than water, and the DNA is dissolved in water, and we always remove it. 2) after extraction of the organic reagents such as phenol and chloroform, it is not possible to put the sampler flat or upside-down to prevent the flow of the liquid from damaging the samplers, and then discard the sample head immediately after adding the sample. 3)? Genome DNA is so large that it can be easily broken by mechanical shear. Therefore, in order to ob