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Primer Combination for Identifying the Authenticity of Chinese Cabbage Varieties

A primer combination and primer set technology, applied in the biological field, can solve problems such as false positives, false negatives, inability to adapt to automation, high-throughput, large-scale, etc.

Active Publication Date: 2021-06-04
BEIJING ACADEMY OF AGRICULTURE & FORESTRY SCIENCES
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

At present, SSR molecular markers are mostly used in other crops, but the SSR detection method can easily lead to false, false positive, and false negative results; it cannot meet the requirements of automation, high throughput, and large scale

Method used

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  • Primer Combination for Identifying the Authenticity of Chinese Cabbage Varieties
  • Primer Combination for Identifying the Authenticity of Chinese Cabbage Varieties
  • Primer Combination for Identifying the Authenticity of Chinese Cabbage Varieties

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0039] Embodiment 1, the acquisition of primer combinations for identifying the authenticity of Chinese cabbage varieties

[0040] 1. Discovery of 32 SNP sites

[0041] The present invention obtains 32 SNP sites based on the resequencing data of 20 Chinese cabbage representative resources. These 20 choy sums represent rich types of resources, covering thick-skinned choy hearts (8 parts), thin-skinned choy hearts (7 parts), wild choy hearts (1 part) and intermediate types (4 parts), basically including the main types of choy hearts Ecotypes, with high genetic diversity in terms of agronomic traits, represent as much germplasm representation as possible.

[0042] Specifically, the screening criteria for SNP loci are as follows: evenly selected positions throughout the genome, good polymorphism, low heterozygosity, MAF>0.3, good PCA clustering effect, high discrimination, and conservative 50bp sequences on both wings (no InDel , no SSR, no other SNP).

[0043] For the basic in...

Embodiment 2

[0056] The validity check of the primer combination that embodiment 2, embodiment 1 develop

[0057] Randomly select 179 Chinese cabbage varieties for testing to check the effectiveness of the primer combinations developed in Example 1.

[0058] The basic information of the 179 Chinese cabbage varieties tested is shown in Table 3. The 179 tested Chinese cabbage varieties are all common fine varieties or some imported varieties.

[0059]Table 3. Basic information of 179 Chinese cabbage varieties tested

[0060]

[0061]

[0062]

[0063] 1. Obtaining the genomic DNA of the tested cabbage varieties

[0064] Genomic DNA of 179 leaves (true leaves of 30 seeds were mixed) were extracted by CTAB method to obtain genomic DNA of tested Chinese cabbage varieties.

[0065] The quality and concentration of the genomic DNA of the tested cabbage varieties must meet the requirements of PCR, and the standard is: agarose electrophoresis shows a single DNA band without obvious dispe...

Embodiment 3

[0077] Embodiment 3, establishing the method for detecting which Chinese cabbage to be tested belongs to among the 179 Chinese cabbage varieties in embodiment 2

[0078] One, establish the method for detecting which kind of Chinese cabbage to be tested belongs to among the 179 Chinese cabbage varieties in embodiment 2

[0079] 1. Obtaining the genomic DNA of Chinese cabbage to be tested

[0080] According to the method of step 1 in the embodiment 2, "leaves of Chinese cabbage varieties to be tested" are replaced with "leaves of Chinese cabbage to be tested", and other steps are all unchanged, and the genomic DNA of Chinese cabbage to be tested is obtained.

[0081] 2. Using the genomic DNA of Chinese cabbage to be tested as a template, 32 primer sets were used for PCR amplification to obtain PCR amplification products. In each PCR reaction system, the concentration ratio of the primers containing "F1" in the name, the primers containing "F2" in the name and the primer contain...

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Abstract

The invention discloses a combination of primers for identifying the authenticity of Chinese cabbage varieties. The primer combination provided by the present invention consists of 32 primer sets. Each primer set consists of 3 primer sequences and is used to amplify a SNP locus. The nucleotide sequences of each primer in the 32 primer sets are shown in sequence from SEQ ID NO:1 to SEQ ID NO:144. The combination of primers provided by the invention can be used for early identification of Chinese cabbage varieties at the seed or seedling stage, to ensure the authenticity of the varieties, to effectively protect the rights and interests of producers and breeders, and to provide technology for the protection of Chinese cabbage germplasm resources and new varieties. support. The method provided by the invention can not only identify unknown cabbage varieties, but also identify the authenticity of known varieties. The method provided by the invention has the advantages of high throughput, accuracy, low cost, simple operation, saving manpower and material resources, etc., and has very broad application prospects.

Description

technical field [0001] The invention belongs to the field of biotechnology, and in particular relates to a combination of primers for identifying the authenticity of Chinese cabbage varieties. The method can be applied to biotechnology promotion services and provide technical support for the protection of Chinese cabbage germplasm resources and varieties. Background technique [0002] Caixin is one of the specialty vegetables in southern my country, also known as cabbage sprouts. Caixin, a subspecies of Brassica cabbage in the family Brassicaceae, is an annual or biennial herb, known as "the queen of vegetables" and "the crown of vegetables". Caixin is a continental vegetable in Guangdong, Guangxi and other places. It can be cultivated all year round in Guangdong, and the perennial sowing area is large. According to statistics from the Guangzhou Municipal Vegetables Office in 2012, the city’s annual planting area of ​​Chinese cabbage reached 252,000 mu, and the sowing area ...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12Q1/6895C12N15/11
CPCC12Q1/6895C12Q2600/156
Inventor 温常龙张德双张建杨静静罗江张晓飞杨明珠
Owner BEIJING ACADEMY OF AGRICULTURE & FORESTRY SCIENCES
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