Efficient extracting method for Wuta-tsai total RNA

An extraction method, the technology of Wutaicai, which is applied in the field of efficient extraction of total RNA from Wutaicai, can solve problems affecting research and achieve the effect of improving RNA purity and saving extraction time

Inactive Publication Date: 2016-05-04
ANHUI AGRICULTURAL UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

[0003] After the cell wall of Wutaia cells is broken, secondary metabolites such as polysaccharides and proteins interfere with the extraction of RNA in d

Method used

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  • Efficient extracting method for Wuta-tsai total RNA
  • Efficient extracting method for Wuta-tsai total RNA

Examples

Experimental program
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Effect test

Embodiment 1

[0031] The high-efficiency extraction method of the total RNA of embodiment 1 Wutah

[0032] 1. Take 0.1 mg of fresh young leaves of Utassica chinensis, cut them into pieces, add 20 mL of liquid nitrogen in 2-3 times, grind the leaves into powder within 3 minutes, pour them into a centrifuge tube, and complete the operation on an ultra-clean workbench ( sheltered environment).

[0033] 2. Add 1mL Trizol lysate to the above powder, place the centrifuge tube in a vortex machine for vigorous shaking, let it stand at room temperature for 3min, and then centrifuge at 12000rpm at 4°C for 5min.

[0034] 3. Put the supernatant in another centrifuge tube, add 0.2mL chloroform to the supernatant, cover the centrifuge tube tightly, shake vigorously by hand for 20s, let stand at room temperature for 5min, then centrifuge at 13000rpm for 10min at 4°C.

[0035] 4. Transfer the supernatant to another centrifuge tube, add an equal volume of isopropanol at 4°C, shake up and down, let stand at...

Embodiment 2

[0037] The high-efficiency extraction method of the total RNA of embodiment 2 Wutah

[0038] 1. Take 0.1 mg of fresh young leaves of Wutaicai, cut them into pieces, add 20 mL of liquid nitrogen in 2-3 times, grind the leaves into powder within 5 minutes, pour them into a centrifuge tube, and complete the operation on an ultra-clean workbench ( sheltered environment).

[0039] 2. Add 1.2mL Trizol lysate to the above powder, place the centrifuge tube in a vortex machine for vigorous shaking, let it stand at room temperature for 4min, and then centrifuge at 12000rpm at 4°C for 4min.

[0040] 3. Put the supernatant in another centrifuge tube, add 0.15mL chloroform to the supernatant, cover the centrifuge tube tightly, shake vigorously by hand for 15s, let it stand at room temperature for 4min, and then centrifuge at 13000rpm for 5min at 4°C.

[0041] 4. Transfer the supernatant to another centrifuge tube, add an equal volume of isopropanol at 4°C, shake up and down, let stand at ...

Embodiment 3

[0043] The high-efficiency extraction method of the total RNA of embodiment 3 Wutah

[0044] 1. Take 0.1 mg of fresh young leaves of Wutaicai, cut them into pieces, add 15 mL of liquid nitrogen in 2-3 times, grind the leaves into powder within 3 minutes, pour them into a centrifuge tube, and complete the operation on an ultra-clean workbench ( sheltered environment).

[0045] 2. Add 1mL Trizol lysate to the above powder, place the centrifuge tube in a vortex machine for vigorous shaking, let it stand at room temperature for 5min, and then centrifuge at 12000rpm at 4°C for 5min.

[0046] 3. Put the supernatant in another centrifuge tube, add 0.3mL chloroform to the supernatant, cover the centrifuge tube tightly, shake vigorously by hand for 30s, let stand at room temperature for 3min, then centrifuge at 13000rpm for 15min at 4°C.

[0047]4. Transfer the supernatant to another centrifuge tube, add an equal volume of isopropanol at 4°C, shake up and down, let stand at -20°C for ...

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Abstract

The invention relates to an efficient extracting method for Wuta-tsai total RNA. The method comprises the steps that liquid nitrogen is added into Wuta-tsai leaves, grinding is carried out, the centrifugal rotating speed is increased, ethyl alcohol of different concentrations is used for washing, and RNA sedimentation is carried out, so that extraction of high-purity, high-concentration and degradation-free Wuta-tsai total RNA is achieved. The method for scientifically and systematically extracting total RNA from the Wuta-tsai leaves is provided for the first time, the extraction efficiency is improved, and the problem that pollution is produced in the extraction process to cause RNA degradation can be effectively avoided. By the adoption of the method, Wuta-tsai total RNA extraction can be completed within only 50-55 min, the high-purity, high-concentration and degradation-free total RNA is obtained, and deepening of follow-up study is facilitated.

Description

technical field [0001] The invention relates to a process for extracting plant RNA, in particular to a method for efficiently extracting the total RNA of Utah cabbage. Background technique [0002] Wutaicai, also known as Dacai, Heicai, etc., is a variety of Brassicaceae Brassica subspecies. It is fed with dark green leaves and is mainly distributed in the Yangtze River Basin of my country. At present, no suitable RNA extraction method has been established for the molecular biology research of Utah. [0003] After the cell wall of Wutaia cells is broken, secondary metabolites such as polysaccharides and proteins interfere with the extraction of RNA in different ways. The RNA enzymes in the cells and in the operating system will degrade the RNA in the isolated tissue, thereby affecting follow up research. On the basis of fully referring to the RNA extraction methods of other cruciferous crops, the present invention establishes a high-efficiency extraction method for the tot...

Claims

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Application Information

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IPC IPC(8): C12N15/10
CPCC12N15/1003C12Q2523/32
Inventor 汪承刚方柔刘思嘉陈国户袁凌云朱世东葛继涛
Owner ANHUI AGRICULTURAL UNIVERSITY
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