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Simple and rapid method for extracting rice DNA

An extraction method and rice technology, which are applied in the field of simple and rapid extraction of rice DNA based on PCR, can solve the problems of difficulty in applying plant leaf genomic DNA extraction, unsuitable sample DNA extraction, laborious process, etc. The effect of simple operation

Inactive Publication Date: 2018-03-23
XUZHOU INST OF AGRI SCI IN JIANGSU XUHUAI DISTRICT (JIANGSU XUZHOU SWEETPOTATO CENT)
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0003] Predecessors have summarized a series of methods for the extraction of plant genomic DNA, such as SDS method and CTAB method. Although the quality of DNA extracted by these methods is good, the process is laborious and cumbersome, requiring the use of toxic chemicals such as chloroform and isoamyl alcohol. Reagents, difficult to apply to the needs of genomic DNA extraction of a large number of plant leaves
Correspondingly, some biological companies have also developed some genomic DNA extraction kits, but the kits are often costly and are not suitable for the extraction of DNA from a large number of samples.

Method used

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  • Simple and rapid method for extracting rice DNA
  • Simple and rapid method for extracting rice DNA

Examples

Experimental program
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Effect test

Embodiment 1

[0021] Example 1: Using primer Pi-hk1-P15 to amplify rice leaf DNA

[0022] On May 17, 2017, the seedlings were raised in the seedling field, transplanted to the field on June 15, and samples were taken after one month. Cut the leaves into 2ml centrifuge tubes, add 500µl extract, treat at 65°C for 30min, and centrifuge at 10,000rpm for 5min , move the supernatant to a 96-well plate filled with 100µl isopropanol, let it stand for 30min, remove the supernatant after centrifugation, add 200µl of 70% absolute ethanol to wash with a row gun, dry it, add 100-200µl to extinguish ddH 2 O or TE solution to dissolve. The extract in this example is: 12.114g Tris (Tris), 3.722g Disodium Ethylenediaminetetraacetate (EDTA), 74.55g Potassium Chloride (KCl), add water to 1L, pH= 8.0.

[0023] Using the DNA extracted by this method as a template, PCR amplification was performed using the gene primer Pi-hk1-P15 (see Table 1 for the sequence). The 20 µl PCR system used was: 1 µl DNA template,...

Embodiment 2

[0025] Example 2: Using SSR primer RM24097 to amplify rice leaf DNA

[0026] On May 17, 2017, the seedlings were grown in the seedling field, and transplanted to the field on June 15. After one month, the samples were taken. Cut the leaves into 2ml centrifuge tubes, add 500µl of extract, and treat at 65°C for 30min, 10000r.min -1 Centrifuge for 10 minutes, transfer the supernatant to a 96-well plate containing 100 µl of isopropanol, let it stand for 30 minutes, remove the supernatant after centrifugation, add 200 µl of 70% absolute ethanol to wash with an exhaust gun, dry it, and add 100-200 µl ddH after sterilization 2 O dissolved. The extract in this example is: 12.114g Tris (Tris), 3.722g Disodium Oxalate Tetraacetate (EDTA), 74.55g Potassium Chloride (KCl), add water to 1L, Ph= 8.0.

[0027] Using the DNA extracted by this method as a template, PCR amplification was performed using the gene primer RM7654 (see Table 1 for the sequence). RM24097 is located on chromosome 9...

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PUM

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Abstract

The invention provides a method for extracting rice DNA. The method comprises the following steps: cutting rice leaves into pieces, placing the rice leaves into a 2ml centrifugal tube, adding an extracting solution, treating the rice leaves in an oven at 65 DEG C for about 30 min, carrying out centrifugation, placing the supernatant in a 96-well plate containing isopropanol, carrying out standingfor about 30 min, and carrying out centrifugation to obtain a white solid, namely DNA. Compared with a traditional DNA extraction technology, the method provided by the invention has the advantages ofsimple and rapid operation, less reagent used, low cost, no need of phenol and chloroform extraction, environment-friendliness and no need of liquid nitrogen or frozen sample. The DNA extracted by the method provided by the invention has the characteristics of high yield, good quality, high success rate of PCR amplification, low cost, good universality, strong molecular stability and the like.

Description

technical field [0001] The invention relates to the field of biotechnology, in particular to a simple and rapid method for extracting rice DNA based on PCR. Background technique [0002] Rice is one of the three most important food crops in the world. It has a rich and in-depth genome research foundation, among which DNA preparation is the basis for in-depth molecular biology research. PCR-based molecular marker technology has been widely used in gene positioning, gene cloning, detection of transgenic plants, and molecular marker-assisted breeding. These studies require a lot of DNA extraction. However, DNA extraction is a tedious work that consumes manpower and material resources. Therefore, it is very necessary to establish a simple, efficient and low-cost DNA extraction method suitable for PCR analysis. [0003] Predecessors have summarized a series of methods for the extraction of plant genomic DNA, such as SDS method and CTAB method. Although the quality of DNA extrac...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/10
CPCC12N15/1003
Inventor 何弯弯王健康丁成伟郭荣良吴玉玲徐家安王友霜胡婷婷赵轶鹏
Owner XUZHOU INST OF AGRI SCI IN JIANGSU XUHUAI DISTRICT (JIANGSU XUZHOU SWEETPOTATO CENT)
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