MNP core primer combination for eggplant DNA variety molecular identification and application thereof

A technology of molecular identification and primer combination of varieties, applied in the field of molecular biology, can solve problems such as hindering the development of MNP sites

Active Publication Date: 2021-03-12
WUHAN ACADEMY OF AGRI SCI +1
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Due to the lack of eggplant resequencing data published in NCBI and other databases, the development of MNP sites is hindered

Method used

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  • MNP core primer combination for eggplant DNA variety molecular identification and application thereof
  • MNP core primer combination for eggplant DNA variety molecular identification and application thereof
  • MNP core primer combination for eggplant DNA variety molecular identification and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0024] Screening and Validation of MNP Core Marker Primer Combinations for Molecular Identification of Eggplant DNA Varieties (1) Screening of Eggplant MNP Primer-Marker Combinations

[0025]Using the publicly released SME_r2.5.1 sequence as the eggplant reference genome (http: / / eggplant.kazusa.or.jp / ), the total length is 850,044,630bp, assembled to the contig level; two varieties provided by Wuhan Academy of Agricultural Sciences (PnP05 and GnP05, species name link https: / / doi.org / 10.1371 / journal.pone.0179491) genome resequencing data, eight NCBI eggplant resequencing data (NCBI accession numbers are SRR5796636 to SRR5796643), and publicly available The NCBI accession numbers of eggplant resequencing data are GB AccessionNo.30075032 to 3007503258; No.30075103 to 30075116. Based on the comparative analysis of the above sequencing data, 522 MNP marker sites were screened, and primers were designed (see Table 1 for primer details).

[0026] (2) Correctness analysis of eggplant...

Embodiment 2

[0032] Construction of Standard DNA Fingerprint of Eggplant by MNP Core Marker Primer Combination for Molecular Identification of Eggplant DNA Varieties

[0033] 1. DNA extraction and quality assessment

[0034] Genomic DNA of all eggplant varieties used to construct the eggplant fingerprint database was extracted by conventional CTAB method, and the quality of DNA was detected by agarose gel and ultraviolet spectrophotometer after RNase treatment. If the DNA band is single, there is no obvious degradation, the UV spectrophotometer detects that the A260 / 280 is between 1.8-2.0, and the A260 / 230 is between 1.8-2.0, there is no obvious light absorption at 270nm, and the DNA concentration is greater than 50ng / ul, it means Genomic DNA meets the relevant quality requirements and can be used for subsequent experiments.

[0035] 2. MNP marker detection

[0036] Using 522 synthetic eggplant MNP-labeled primers, 80 eggplant inbred varieties provided by the Vegetable Research Institute...

Embodiment 3

[0040] Authenticity of eggplant varieties detected by MNP core marker primer combination for molecular identification of eggplant DNA varieties

[0041] 1. Determination of the differentiation degree of eggplant MNP primer marker varieties and the determination of the authenticity test results of eggplant varieties

[0042] The 80 eggplant inbred line samples in Example 2 were pairwise compared to analyze the differences of their MNP marker sites, and a total of 3160 pairs of comparison results were obtained. The average number of MNP marker differences between any varieties was 226.79, and the genetics of 3160 pairs of samples The difference in distance is between 0% and 83.69%, with an average difference of 44.16%, among which 8 pairs have a difference of 0, accounting for 0.25% of the total number of sample pairs. Using the developed marker detection method, 99.75% of eggplant varieties could be distinguished ( figure 2 ), indicating that the developed eggplant MNP marker...

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Abstract

The invention belongs to the technical field of molecular biology, and particularly discloses an MNP core primer combination for eggplant DNA variety molecular identification and application thereof.The primer combination is shown by a table 1. The primer combination composed of 522 MNP core primers provided by the invention can be compatible with a next-generation sequencing platform, has the characteristics of high efficiency, correctness, high reproducibility and the like, and meets the requirements of eggplant standard fingerprint data construction; the authenticity of the eggplant sampleto be detected can be accurately identified by identifying the eggplant sample to be detected and the control variety; analysis on eggplant high-generation selfing line materials shows that the MNP site has high polymorphism and repeatability, can distinguish different eggplant groups, and has a wide application range.

Description

technical field [0001] The invention belongs to the technical field of molecular biology, and in particular relates to a combination of MNP core primers for molecular identification of eggplant DNA varieties and an application thereof. [0002] technical background [0003] Eggplant (Solanum melongena L.) is a vegetable crop of the genus Solanaceae, which is widely cultivated all over the world. It is the fourth largest vegetable crop in the world and one of the main vegetable crops in my country. According to the 2018 statistical report of the Food and Agriculture Organization of the United Nations (FAO), the global eggplant production is about 50 million tons, of which Asian eggplant accounts for more than 90% of the total production. More than 50% of the output. my country is the secondary origin of eggplant, with a long history of cultivation and abundant germplasm resources. The National Germplasm Medium-term Bank has collected 1468 germplasms. Crop germplasm resources...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/6895C12Q1/6806C40B50/06C12N15/11G16B20/40G16B25/20G16B30/10G16B40/10G16B40/30G16B45/00G16B50/00
CPCC12Q1/6806C12Q1/6895C12Q2600/156C40B50/06G16B20/40G16B25/20G16B30/10G16B40/10G16B40/30G16B45/00G16B50/00C12Q2525/191C12Q2531/113
Inventor 张洪源张敏谈杰黄树苹陈霞
Owner WUHAN ACADEMY OF AGRI SCI
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