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A functional marker wx-a/b for analyzing rice wx gene using DNA melting temperature and its use method and application

A technology of melting temperature and wx-a, which is applied in the direction of DNA/RNA fragments, recombinant DNA technology, microbial measurement/inspection, etc., can solve the problems of restricting wide application and involving many links, achieve good repeatability and avoid gelation The effect of electrophoresis and fully closed genotype detection

Active Publication Date: 2015-09-30
SOUTH CHINA AGRI UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The use of DNA melting temperature to determine genotypes has been reported in the study of human diseases. However, this technology involves many links and requires detailed pre-optimization work. At present, there are few reports on the systematic research of this technology on rice. To some extent, it limits its wide application in rice breeding

Method used

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  • A functional marker wx-a/b for analyzing rice wx gene using DNA melting temperature and its use method and application
  • A functional marker wx-a/b for analyzing rice wx gene using DNA melting temperature and its use method and application
  • A functional marker wx-a/b for analyzing rice wx gene using DNA melting temperature and its use method and application

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0032] Example 1: Development of a functional marker for rice Wx gene analysis using DNA melting temperature.

[0033] 1. Extraction of genomic DNA of rice varieties R173 and Nipponbare.

[0034]Concrete method: 1) take a small amount of paddy rice seedling transplanting one month young leaves and place in liquid nitrogen freezing 2.0mL sterilized centrifuge tube and stir until powdery, add 1000 μ L 2 × CTAB-DNA extraction solution (mass fraction W / V 2% CTAB , pH8.0; mass fraction W / V 1% of PVP; 100mmol / L Tris-HCl, pH8.0; 1.4mol / L NaCl; 20mmol / L EDTA, pH8.0; volume fraction V / V 0.2% of mercapto ethanol); 2) Place it in a constant temperature water bath at 65°C and shake it every 10 minutes, and take it out after 30-45 minutes; 3) After cooling for 2 minutes, add 1000 μL chloroform-isoamyl alcohol (volume ratio 24:1), shake vigorously up and down, Mix the two evenly; 4) Centrifuge at 10000rpm for 10min, gently remove the supernatant into a 1.5ml sterilized new centrifuge tube,...

Embodiment 2

[0048] Example 2: Using Wx-a / b to detect rice Wx genotype.

[0049] 1. Test materials.

[0050] The research materials in this experiment are indica rice variety R173 and japonica rice variety Nipponbare.

[0051] 2.2 Identification of the amylose content of the test materials.

[0052] Amylose content identification method is the same as described in Example 1.

[0053] The results showed that R173 was a variety with high amylose content (26.5%), and Nipponbare was a variety with low amylose content (12.3%).

[0054] 3.2 DNA extraction from the test materials, PCR amplification analysis.

[0055] DNA extraction and PCR amplification methods are the same as those described in Example 1.

[0056] 4. The detection method of the DNA melting temperature of the amplified product of functionally labeled Wx-a / b.

[0057] The melting temperature detection method is as described in Example 1.

[0058] The results showed that the DNA melting temperature of the R173 amplified produ...

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Abstract

The invention relates to the technical field of biology, and specifically discloses a functional marker Wx-a / b for analyzing rice Wx gene by using a DNA melting temperature, a use method and an application thereof. The functional marker Wx-a / b comprises primers Wx-a / b-out and Wx-a / b-in, and can be provided for detecting Wx genotype, wherein genotype detection with characteristics of batch, automation and full closing can be achieved with the functional marker Wx-a / b. The detection method has characteristics of low cost, simpleness, good practicality and good repeatability.

Description

technical field [0001] The invention relates to the field of biotechnology, more specifically, to a method for analyzing water by using DNA melting temperature [0002] Functional marker Wx-a / b of rice Wx gene and method and application thereof. Background technique [0003] Rice is an important crop in my country, and moderate amylose content is an important characteristic of high-quality rice. The amylose content (Amylose Content, AC) of rice is one of the most important factors affecting the taste of rice; indica rice with medium and low AC is favored by consumers because it is soft but not too sticky. The traditional amylose content selection method is determined by chemical method, which has the disadvantages of destructiveness and low efficiency; at the same time, chemical method cannot be measured in the low generation of breeding (such as F2 single plant generation). The most effective selection should be to select directly on the amylose genotype. [0004] Studie...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12Q1/68C12N15/11
Inventor 郭涛罗文龙陈志强王慧周丹华刘永柱张建国
Owner SOUTH CHINA AGRI UNIV