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Functional marker Wx-a/b for analyzing rice Wx gene by using DNA melting temperature, use method and application thereof

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 problems involving many links and limit wide application, and avoid gel electrophoresis, repeatability, etc. Good, fully closed genotype detection effect

Active Publication Date: 2013-12-11
SOUTH CHINA AGRI UNIV
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  • 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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  • Functional marker Wx-a/b for analyzing rice Wx gene by using DNA melting temperature, use method and application thereof
  • Functional marker Wx-a/b for analyzing rice Wx gene by using DNA melting temperature, use method and application thereof
  • Functional marker Wx-a/b for analyzing rice Wx gene by using DNA melting temperature, use method and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0033] Embodiment 1: A kind of utilization DNA melting temperature analysis rice wxya Functional marker development of genes.

[0034] 1. Extraction of genomic DNA of rice varieties R173 and Nipponbare.

[0035] Specific methods: 1) Take a small amount of young rice leaves one month after transplanting, put them in a 2.0 mL sterilized centrifuge tube frozen in liquid nitrogen, stir until powdery, add 1000 μL 2×CTAB-DNA extract (mass fraction W / V 2% CTAB , pH8.0; mass fraction W / V 1% of PVP; 100 mmol / L Tris-HCl, pH8.0; 1.4 mol / L NaCl; 20 mmol / L EDTA, pH8.0; volume fraction V / V 0.2% mercaptoethanol); 2) placed in a constant temperature water bath at 65 °C and shaken every 10 min, and removed after 30-45 min; 3) after cooling for 2 min, add 1000 μL chloroform-isoamyl alcohol (volume ratio 24:1) , shake vigorously up and down to mix the two evenly; 4) Centrifuge at 10,000 rpm for 10 min, gently remove the supernatant into a 1.5 ml sterilized new centrifuge tube, add 600 μL of pr...

Embodiment 2

[0049] Embodiment 2: Utilize Wx-a / b to detect rice wxya genotype.

[0050] 1. Test materials.

[0051] The research materials in this experiment are indica rice variety R173 and japonica rice variety Nipponbare.

[0052] 2. Identification of amylose content in 2 test materials.

[0053] Amylose content identification method is the same as described in Example 1.

[0054] 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%).

[0055] 3. 2 copies of test material DNA extraction, PCR amplification analysis.

[0056] DNA extraction and PCR amplification methods are the same as those described in Example 1.

[0057] 4. Detection method of DNA melting temperature of functionally labeled Wx-a / b amplified product.

[0058] The melting temperature detection method is as described in Example 1.

[0059] The results showed that the melting temperature of R173 amplification product DNA was 1°C hi...

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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

[0001] technical field [0002] The invention relates to the field of biotechnology, more specifically, to a method for analyzing water by using DNA melting temperature [0003] rice wxya Functional marker Wx-a / b of gene and its use method and application. Background technique [0004] 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 popular with consumers because it is soft but not too sticky. The traditional selection method of amylose content 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....

Claims

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Application Information

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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