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Single nucleotide polymorphism marker site, primers and reagent kit for identifying peach fruit flesh color characteristic and application

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

Active Publication Date: 2016-10-26
ZHENGZHOU FRUIT RES INST CHINESE ACADEMY OF AGRI SCI
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  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0005] However, the existing SSR markers linked to peach fruit flesh color (white / yellow) traits 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 linked to target traits Mostly from the results of microarray identification, due to the small number of microarray sites (less than 9000), it cannot be guaranteed that the identified SNPs are the most associated or linked loci with the target traits

Method used

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  • Single nucleotide polymorphism marker site, primers and reagent kit for identifying peach fruit flesh color characteristic and application
  • Single nucleotide polymorphism marker site, primers and reagent kit for identifying peach fruit flesh color characteristic and application
  • Single nucleotide polymorphism marker site, primers and reagent kit for identifying peach fruit flesh color characteristic and application

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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 flesh color were located at 24827153 bp of chromosome 1.

Embodiment 2

[0028] Embodiment 2 Utilizes the method for identifying peach fruit flesh color (white / yellow) traits with 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 24827153rd position of the first chromosome 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 biotec...

Embodiment 3

[0063] Example 3 Blind test verification of phenotypic traits in 15 hybrid populations using fruit flesh color-associated SNP markers

[0064] 1. Selection of experimental materials

[0065] 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.

[0066] Table 6 The name and population size information of the tested hybrid population

[0067]

[0068]

[0069] 2. Identification method using fruit flesh color-associated SNP markers

[0070] Using the 24827153rd position of the first chromosome of the peach genome of the present invention as the nucleotide polymorphism marker site, a total of 221 peach individual plant fruit flesh colors of 15 hybrid populations were blindly tested and identified. For the specific identification method,...

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Abstract

The invention discloses a single nucleotide polymorphism marker site, primers and a reagent kit for identifying a peach fruit flesh color characteristic and application. The single nucleotide polymorphism marker site is the 24827153th nucleotide of the first chromosom of peach genomes, and the nucleotide is A or G. The screened 24827153th nucleotide of the first chromosom of peach genomes serves as the single nucleotide polymorphism marker site, can be used for identifying or assisting in identifying the peach fruit flesh color characteristic and is high in preparation rate when used for identifying the peach fruit flesh color characteristic. The SNPs accuracy rates of 221 parts of peach single plants to be detected in 15 hybrid groups are verified, and results show that the average accuracy rate of the fruit flesh color SNPs is increased to 86.43% from 73.76% compared with other lately-reported research. It means that the single nucleotide polymorphism marker site has the advantages of being simple, fast and low in cost when used for detection, and large-scale application in production can be achieved.

Description

technical field [0001] The invention relates to a single nucleotide polymorphism marker site, a primer, a kit and an application for identifying peach fruit flesh color (white / yellow) traits, 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 relati...

Claims

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Application Information

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IPC IPC(8): C12Q1/68C12N15/11
Inventor 王力荣曹珂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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