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Method for extracting plant DNA and RNA at the same time

A plant and extraction technology, which is applied in chemical instruments and methods, plant raw materials, plant/algae/fungus/moss components, etc., can solve the problems of high cost, low extraction purity, and affecting the purity of nucleic acid, so as to save time and extract high quality effect

Inactive Publication Date: 2009-07-22
ZHEJIANG ACADEMY OF AGRICULTURE SCIENCES
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  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

However, the extraction purity is relatively low, it is difficult to dissolve (it needs to be dissolved in 8mM NaOH solution), and the DNA fragment is severely broken, only about 10Kb, so it is difficult to meet the downstream experimental requirements
Yam tubers are rich in polysaccharides, which can easily cause co-precipitation during nucleic acid extraction, thus affecting the purity of nucleic acids
At present, in order to remove the interference of polysaccharides at home and abroad, the classic CsCl ultracentrifugation method is used, but this method requires high technical requirements and high cost

Method used

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  • Method for extracting plant DNA and RNA at the same time
  • Method for extracting plant DNA and RNA at the same time

Examples

Experimental program
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Effect test

Embodiment 1

[0036] 1. Take yam tubers, cut them into small pieces with a knife, take 1 gram, and grind them into fine powder with liquid nitrogen according to the ratio of sample: insoluble PVP (K40) = 10:1.

[0037] 2. Add 2.5mL of preheated 2% CTAB extract, and add β-mercaptoethanol to a final concentration of 5%, bathe in water at 65°C for 30 minutes, and shake 3-4 times during the period.

[0038] 3. Add 625uL of absolute ethanol and 275uL of KAc buffer (pH5.3), shake vigorously and mix well, then place in a water bath at 65°C for 10 minutes, shaking 1-2 times during the period.

[0039] 4. Add an equal volume of 3.4 mL of chloroform: isoamyl alcohol (volume ratio 24:1), invert up and down for 5 minutes to mix well, and then centrifuge at 4°C and 16,000 rpm for 10 minutes

[0040]5. Aspirate 700uL of the supernatant into a new 1.5mL eppendorf tube (each tube of the above-mentioned centrifuge tube is divided into two tubes), add 350uL of chloroform:isoamyl alcohol (24:1) and 350uL of w...

Embodiment 2

[0056] Take 1 gram each of the leaves of yam, bamboo, eggplant and gourd, chop them with a knife, and grind them into fine powder with liquid nitrogen according to the ratio of sample:insoluble PVP=10:1. Other steps are with embodiment 1. The obtained RNA and DNA quality are the same as example 1, the RNA ratio is between 1.8~2.0, the DNA ratio is between 1.7~18, and the electrophoresis result band is clear and complete (see Figure 1~2 ).

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Abstract

The invention provides a method for extracting DNA and RNA of a plant synchronously, which utilizes one plant sample; and after the plant sample is treated and decontaminated with the same steps, the DNA and RNA thereof are settled and purified respectively, thereby achieving the purpose of extracting synchronously. The method can be applied to the synchronous extraction of DNA and RNA of a yam sample and has the advantages of being able to synchronously conduct nucleic acid extraction, time-saving, economical and fast; besides, the ultracentrifugation, KAC and the like adopted by the method are very effective in eliminating the impurities contained in the yam such as amylose and the like; the DNA / RNA extraction quality of the method is high; the RNA OD260 / 280 ratio is between (1.8 to 2.0) and the DNA is between 1.7 to 1.8; and the electrophoretic band is complete and clear. The method is also applicable to the nucleic acid extraction of other plant samples.

Description

(1) Technical field [0001] The invention relates to a method for simultaneously extracting plant sample DNA and RNA, in particular to a method for simultaneously extracting DNA and RNA from yam. (2) Background technology [0002] In the past ten years, molecular biology has developed rapidly, and has been widely used in various fields such as medicine, agriculture, pharmacy, forensic science, etc., which has important development significance. Since the discovery of the double helix structure of DNA, nucleic acid has become the focus of biological research, and the discipline of molecular biology with DNA and RNA as the main research objects has emerged. Isolating high-purity and high-quality nucleic acids from samples is the most basic prerequisite for the smooth progress of downstream experiments. DNA and RNA purification techniques mainly include phenol chloroform extraction, ion exchange, salting out, glass milk method and silica gel column method, etc., but these metho...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07H21/02C07H21/04C07H1/08A61K36/8945A61K36/899A61K36/81
Inventor 王玲平周生茂戴丹丽曹家树杨悦俭
Owner ZHEJIANG ACADEMY OF AGRICULTURE SCIENCES
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