Mononucleotide polymorphic marker site, primers and kit for identifying peach fruit epidermal hair characters, and application thereof

A technology of single nucleotide polymorphism and marker site, which is applied in the determination/inspection of microorganisms, biochemical equipment and methods, DNA/RNA fragments, etc. SNPs and other problems, to achieve the effect of low cost and high accuracy

Active Publication Date: 2016-12-07
ZHENGZHOU FRUIT RES INST CHINESE ACADEMY OF AGRI SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0005] However, the existing SSR markers linked to the traits of peach fruit epidermis are far away from the target gene, and the accuracy rate in the early identification of hybrid offspring is low; while the existing SNPs markers lin

Method used

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  • Mononucleotide polymorphic marker site, primers and kit for identifying peach fruit epidermal hair characters, and application thereof
  • Mononucleotide polymorphic marker site, primers and kit for identifying peach fruit epidermal hair characters, and application thereof
  • Mononucleotide polymorphic marker site, primers and kit for identifying peach fruit epidermal hair characters, and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0026] Example 1 Obtaining of SNPs marker sites

[0027] In the present invention, 129 peach germplasms randomly obtained from the peach germplasm resource garden of Zhengzhou Institute of Pomology, Chinese Academy of Agricultural Sciences were used as samples, and the sample DNA was extracted by conventional CTAB method, and the 129 peach germplasms were reconstructed by Illumina HiSeq 2000 sequencer. Sequencing obtained 121Gb data, covering an average of 89.28% of the peach genome, and the average sequencing depth was about 4.21×. According to the 50-150bp reads obtained by sequencing, it is compatible with the peach reference genome v.1.0 ( http: / / www.rosaceae.org / node / 355 ) were compared, and 4,063,377 SNPs were identified. Using these SNPs to conduct genome-wide association analysis on the phenotypic traits of 129 germplasms, it was identified that the SNPs that were significantly associated with peach fruit fur were located at 17,576,893 bp on chromosome 5.

Embodiment 2

[0028] Example 2 The method for identifying phenotypic traits using SNP markers

[0029] 1. DNA extraction

[0030] The DNA of the peach sample tissue to be tested was extracted by the conventional CTAB method, and the RNA was removed. The total volume of the DNA sample was not less than 15 μl. Measure the OD value of the DNA sample at 260nm and 280nm with a UV photometer, and calculate the DNA content and OD 260 / 280 ratio. DNA sample purity OD 260 / 280 The value should be between 1.8-2.0 and the concentration should be diluted to 10ng / μl.

[0031] 2. Design primers

[0032] Primers were designed according to the 200 bp sequences on the left and right of the 17576893rd position of chromosome 5 of the peach genome (see Table 1 for the specific nucleotide sequence).

[0033] Table 1 SNPs flanking sequence information

[0034]

[0035] Among them, R represents A or G.

[0036] After the primers were synthesized by the biotechnology company, the upstream and downstream pr...

Embodiment 3

[0062] Example 3 Blind test verification of phenotypic traits in 15 hybrid populations using fruit fur associated SNP markers

[0063] 1. Selection of experimental materials

[0064] Taking the conventional peach varieties planted in the resource garden of Zhengzhou Pomology Research Institute as experimental materials, 15 hybrid populations with investigated phenotypic traits were selected from which a total of 221 individual plants were selected. See Table 6 for details.

[0065] Table 6 The name and population size information of the tested hybrid population

[0066]

[0067]

[0068] 2. Identification method using fruit coat-associated SNP markers

[0069] Using the 17576893rd position of the 5th chromosome of the peach genome of the present invention as a nucleotide polymorphism marker site, a total of 221 peach single-plant fruit skins from 15 hybrid populations were blindly tested and identified. For the specific identification method, refer to Example 2. method...

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Abstract

The invention discloses a mononucleotide polymorphic marker site, primers and a kit for identifying peach fruit epidermal hair characters, and application thereof. The mononucleotide polymorphic marker site is 17576893th nucleotide of chromosome 5 of the peach genome, and the nucleotide is A or G. The 17576893th nucleotide mononucleotide polymorphic marker site of chromosome 5 of the peach genome can be used for identification or auxiliary identification on peach fruit epidermal hair characters, and has higher accuracy in the peach fruit epidermal hair character identification process. When the SNPs accuracy of 221 peach individuals to be detected in 15 hybrid populations is verified, the result indicates that the detection accuracy for peach fruit epidermal hairy character can reach 96% or above, the detection accuracy for peach fruit hairless character can reach 80% or above, and thus, the mononucleotide polymorphic marker site has higher accuracy. Thus, the mononucleotide marker site for detection is simple, has the advantages of high speed and low cost, and can implement large-scale application in production.

Description

technical field [0001] The invention relates to a single nucleotide polymorphism marker site, a primer, a kit and an application for identifying the traits of peach fruit epidermis, and belongs to the field of biotechnology. Background technique [0002] Selection is one of the most important links in breeding. It refers to selecting genotypes that meet the requirements in a population for subsequent breeding. However, in traditional breeding, because it is difficult to know the genotype of the offspring, the basis of selection is usually phenotype rather than genotype. This selection method is generally effective for qualitative traits, but for quantitative traits, because The lack of a clear correspondence between its phenotype and genotype makes it inefficient. In addition, for fruit trees with fruit traits as the target, these traits have their specific expression period, usually need to spend 3-5 years or even longer childhood, so the selection time is relatively late....

Claims

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Application Information

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IPC IPC(8): C12Q1/68C12N15/11
CPCC12Q1/6895C12Q2600/13C12Q2600/156
Inventor 曹珂王力荣朱更瑞方伟超陈昌文王新卫王琪
Owner ZHENGZHOU FRUIT RES INST CHINESE ACADEMY OF AGRI SCI
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