Extraction method of mangrove plant DNA

Abstract

The invention relates to an extraction method of mangrove plant DNA, comprising the following steps: (1) taking a leaf to grind, adding preheated 2% of CTAB extract to dissolve, and then adding 2 % of PVP by volume; (2) after water bath, adding a small quantity of RNase for heat preservation, then adding chloroform-isoamylol mixed liquor, and carrying out high-speed centrifugation at normal temperature; (3) putting supernate into a centrifuge tube, adding the preheated 2% of CTAB extract, adding chloroform-isoamylol mixed liquor, and mixing and carrying out high-speed centrifugation at normal temperature; (4) taking supernate and then putting into the centrifuge tube, adding NaCl solution and cold isopropanol, mixing, and putting and precipitating DNA at low temperature; (5) carrying out high-speed centrifugation at low temperature, abandoning the supernate, recovering the precipitate, washing by ethanol, air-drying, adding TE buffer solution, and storing at low temperature. The invention can effectively remove polysaccharide and tannin produced in the DNA extraction process, and the obtained mangrove plant has high DNA content and high purity.

Description

technical field [0001] The invention relates to a plant DNA extraction method, in particular to a DNA extraction method for mangrove plants rich in polysaccharides and tannic acid. Background technique [0002] Mangrove is a rare woody viviparous plant that grows in tidal flats and shoals at the junction of tropical and subtropical land and sea. It is a special ecosystem that transitions from land to sea. There are 23 families, 34 genera, and 81 species of mangrove plants in the world. There are 13 families and 24 species of mangrove trees in my country, mainly distributed in Hainan, Guangdong and the coastal areas of Fujian. Because mangroves grow at the junction of land and sea, they are natural barriers for the land, which is of great significance to reducing the damage caused by natural disasters such as typhoons; the mangrove ecosystem also has the function of absorbing and accumulating heavy metal elements, removing organochlorine pesticides, and improving environmenta...

AI Technical Summary

Problems solved by technology

However, due to the particularity of mangrove plants, there are also some difficulties in the research at the molecular level, one of which is the DNA extraction of mangrove plants, because: mangrove plants are famous for their rich tannins, and their cell walls contain A large number of substances such as tannins and polysaccharides, in the process of nucleic acid extraction, the existence of a large amount of tannins and polysaccharides makes them form co-precipitation with DNA, thus affecting the purity of nucleic acids, although the DNA extraction methods of general land plants are very mature , such as the common classic CTAB method, SDS method, etc., and these methods cannot deal with the problem of polysaccharides well, so that relatively pure DNA cannot be obtained. At present, the most commonly used method for removing polysaccharides at home and abroad is to use the classic CsCl Ultracentrifugation method, but this method has high requirements for experimental equipment and operating technology, and is expensive. In addition, some researchers use purification methods such as column filtration. Although genomic DNA is purified well, it is easy to The loss and breakage of genomic DNA cannot meet the high requirements of DNA operations such as enzyme digestion and connection. In addition, there are few reports on how to remove tannins during the DNA extraction process; if further development of population genetics, systematic Evolution, molecular marker-assisted breeding, etc., due to the large number of experimental samples that need to be processed, the cumbersome and expensive defects of the above-mentioned physical or chemical methods are even more unfavorable for experimental research, and DNA has high requirements. Operations such as molecular cloning and genetic modification cannot be carried out smoothly

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