Primer system used for sweet potato paternity test as well as screening method and kit of primer system

A technology for parentage identification and screening methods, applied in the field of plant species identification, can solve the problems of declining genetic diversity, narrow genetic basis, and single breeding background, and achieve the effects of high polymorphism, high genetic stability, and broad application prospects.

Pending Publication Date: 2017-01-04
CROP RES INST GUANGDONG ACAD OF AGRI SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0002] Since 1949, my country's sweet potato science and technology has developed rapidly. However, in sweet potato breeding, due to the long-term artificial selection of a few traits such

Method used

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  • Primer system used for sweet potato paternity test as well as screening method and kit of primer system
  • Primer system used for sweet potato paternity test as well as screening method and kit of primer system
  • Primer system used for sweet potato paternity test as well as screening method and kit of primer system

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0056] 1) Molecular marker design: Use Primer Premier software to design SSR molecular marker primers according to the conserved regions at both ends of the SSR site. Primer design principles: Tm value is 50-60°C; amplified product size is 100-400bp; primer length is 18-24bp, Amplification rate greater than 80%;

[0057] 2) DNA extraction: collect the young leaves of the test material, use the CTAB method to extract genomic DNA, use 0.7% agarose gel electrophoresis to detect its purity, and ultra-micro spectrophotometer to accurately measure the DNA sample concentration, dilute to 100ng / μL, Store at -20°C for later use;

[0058] 3) PCR amplification: use the genomic DNA obtained in step 2) as a template, and use the primers obtained in step 1) to perform PCR amplification; PCR amplification system 10 μL, as follows: 0.15 μL (2.5U / μL) TaqDNA polymerase, upstream Primer 0.3μL (20pmol / μL), downstream primer 0.3μL (20pmol / μL), 0.8μL dNTPs (2.5mmol / L), 1.0μL genomic DNA (20ng / μL),...

Embodiment 2

[0070] Adopt Puning vegetable species (P1) and Tainong 27 (P2) and its 238 (numbering 1-238) F1 progeny populations to detect the genetic stability of 22 candidate primers screened out in Example 1, that is, molecular markers in the parent generation Whether it conforms to the Mendelian law of inheritance with the offspring. PCR products were separated by polyacrylamide gel electrophoresis.

[0071] Among the 22 pairs of candidate primers, 4 pairs of primers showed contradictory allelic loci between parents and offspring, and the remaining 18 pairs of primers completely conformed to the law of Mendelian inheritance. If the band is not clear, it is easy to make wrong interpretations in the genetic analysis of parents and offspring, and it is not an ideal primer for paternity testing. So finally screened 14 pairs of primers as the primer system for paternity identification of sweet potato. The names of these 14 pairs of primers are as follows: GDAAS0338, GDAAS0354, GDAAS0360, ...

Embodiment 3

[0073] This unit undertakes the job of breeding posts in the southern area of ​​the national sweet potato industrial technology system, and keeps more than 100 commonly used hybrid parents, mainly based on the fingerprint data of 105 commonly used hybrid parents, combined with the 41 resources in Example 1 and the resources in Example 2 240 parent-child materials of the whole sibling group, statistics of individual identification rate DP, cumulative individual identification rate CDP, triplet non-father exclusion rate PE (one parent is known, another parent is known) and triplet accumulation of 14 paternity identification loci Non-parent exclusion rate CPE. The calculation reference formulas of DP, CDP, PE and CPE are as follows:

[0074] Individual recognition rate

[0075] D P = 1 - Σ i = 1 n P i 2 ;

[0076] Cumulati...

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Abstract

The invention discloses a primer system used for a sweet potato paternity test as well as a screening method and a kit of the primer system. Specifically, the screening method comprises the following steps: (1) SSR molecular marker design; (2) DNA extraction; (3) PCR amplification and (4) electrophoretic separation; at last, choosing a primer pair having high polymorphism, good repeated stability and hereditary stability, and a clear amplification strip to make up the primer system for a sweet potato paternity test. The primer system locus screened out by the method has the advantages of high co-dominance and polymorphism, good repeatability, clear strip, high hereditary stability and the like. By taking the advantages of a polymorphic primer with a sweet potato microsatellite molecular marker to conduct the paternity test for sweet potatoes, the method has a wide application prospect in fields such as the judgment and discrimination of genetic relationship of the sweet potatoes, the protection of the genetic relationship, property right dispute and the like.

Description

technical field [0001] The invention relates to a primer system for paternity identification of sweet potato, a screening method and a kit thereof, and belongs to the technical field of plant species identification. Background technique [0002] Since 1949, my country's sweet potato science and technology has developed rapidly. However, in sweet potato breeding, due to the long-term artificial selection of a few traits such as yield and resistance, and the selection of varieties with only a few parents, the breeding background is single and the genetic basis is increasingly narrow. , genetic diversity decline. Therefore, it is necessary to study the genetic diversity of sweet potato germplasm resources in order to fully excavate valuable materials and serve sweet potato genetic breeding. [0003] Parentage identification refers to the application of theories and techniques of medicine, genetics and biology to detect and analyze the genetic relationship between parents and ch...

Claims

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Application Information

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IPC IPC(8): C12Q1/68C12N15/11
CPCC12Q1/6811C12Q1/6895C12Q2600/156C12Q2531/113C12Q2565/125
Inventor 罗忠霞房伯平王章英黄立飞陈新亮陈景益张雄坚姚祝芳
Owner CROP RES INST GUANGDONG ACAD OF AGRI SCI
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