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A SNP marker for identifying pear stone size based on high-resolution melting curves and its application

A high-resolution, melting curve technology, applied in the field of SNP markers, can solve the problems of the development and application of SNP molecular markers that have not yet been reported, and achieve good application value.

Active Publication Date: 2021-09-21
NANJING AGRICULTURAL UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] At present, there are few studies on pear pit size, and there is no report on the development and application of SNP molecular markers for pear pit size

Method used

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  • A SNP marker for identifying pear stone size based on high-resolution melting curves and its application
  • A SNP marker for identifying pear stone size based on high-resolution melting curves and its application
  • A SNP marker for identifying pear stone size based on high-resolution melting curves and its application

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0022] The mensuration of embodiment 1 fruit stone size

[0023] The size of the pear stone is judged by measuring its vertical and horizontal diameters. First, we cut the pear fruit longitudinally, and then use a vernier caliper to measure the core. Take 3-6 fruits of the same variety for repeated measurement, and record the data. The final data are presented as mean ± standard deviation. See Table 1 for the vertical and horizontal diameters of the mature fruits of 72 pear natural populations.

[0024] Table 1 The stone size of mature fruits of 72 pear natural populations

[0025]

[0026]

[0027]

[0028] Note: Serial numbers 1-36 are varieties with small stones, and numbers 37-72 are varieties with large stones.

Embodiment 2

[0029] Example 2 Development of HRM-specific primers using the SNP marker site Pfs258-07

[0030] The DNA sequence of the SNP marker Pfs258-07 on the seventh chromosome was searched from the Dangshan Suli genome database, and the 300 bp sequence before and after this site was selected, and the SNP marker primers were developed and designed according to the primer design principles (Table 2). Using the designed SNP marker primers to carry out PCR amplification on the genomic DNA of Dangshansu pear, the primers were amplified normally, and the PCR product was in line with the predicted size.

[0031] Table 2 SNP marker primers

[0032]

Embodiment 3

[0033] Embodiment 3 Utilizes HRM (high-resolution melting, high-resolution melting curve) technology to carry out classification to pear fruit stone size

[0034] The HRM reaction system was performed according to the instructions in the LightCycler 480 High Resolution Melting Master kit, and the HRM analysis was performed on a LightCycler480II fluorescent quantitative PCR instrument. 20μL reaction system: containing 30ng pear genomic DNA template, 1×Master Mix, 2.5mmol / L MgCl 2 , 200nM of the primer described in claim 1 or 2; the amplification program adopts drop-down PCR (touchdown PCR): pre-denaturation at 95°C for 10min, then denaturation at 95°C for 10s, annealing at 65-53°C (decreasing by 0.5°C per cycle) for 10s, The 10 s extension program at 72°C was performed for 45 cycles. Melting was performed after the PCR cycle was completed. The program was: 95°C for 1 min, 40°C for 1 min, 65°C for 1 s, and then the temperature was continuously raised from 70°C to 95°C. Fluoresc...

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Abstract

The invention discloses a SNP molecular marker primer closely related to the size of pear stones and an application thereof. A SNP marker Pfs258‑07 closely related to pear stone size, primers: SEQ ID NO.1; SEQ ID NO.2. The pear stone size can be well typed based on the high-resolution melting curve with this specific primer. The pear pit size molecular marker primers provided by the invention can be used for molecular marker-assisted selection breeding of pear pit size, and have important theoretical and practical guiding significance for accelerating the genetic improvement process of pear varieties and improving the efficiency of breeding selection.

Description

technical field [0001] The invention belongs to the field of molecular genetic breeding, and relates to a SNP marker for identifying pear stone size based on a high-resolution melting curve and an application thereof. Background technique [0002] Pear fruit not only has the characteristics of juicy flavor, sweet and sour taste, but also is rich in dietary fiber and various nutrients. Pear is the main cultivated fruit tree species in the world, and it is cultivated in 87 countries on five continents. Pear is also the third largest fruit tree species in my country. The cultivated area and output of pears in my country rank first in the world. The eating and appearance quality of pear fruit has always been a concern of people. It is an important factor determining the economic value of pear fruit. Therefore, it is of great significance to improve the eating and appearance quality of pear fruit. The eating and appearance quality of pear fruit are affected by many factors, am...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12Q1/6895C12N15/11
CPCC12Q1/6895C12Q2600/13C12Q2600/156
Inventor 吴俊李晓龙汤子凯张绍铃张询张明月李甲明孙满意齐开杰
Owner NANJING AGRICULTURAL UNIVERSITY
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