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SNP locus combination for detecting resistance of tomato gray leaf spot disease and application thereof

A tomato gray leaf spot and site technology, applied in the field of plant biology, can solve problems such as linkage burden, insufficient versatility of SNP sites, and invalid selection of target genes, to achieve accurate detection, reduce field transplanting workload, and improve The effect of breeding efficiency

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

AI Technical Summary

Problems solved by technology

Through the analysis and identification of a large number of variant group data directly in the adjacent regions of the upstream and downstream of the known Sm gene and within the gene, multiple SNP sites that can quickly and effectively detect the resistance of tomato gray leaf spot can be obtained, which can realize accurate identification of the target gene region and selection, so as to solve the lack of versatility of SNP sites in the prior art, or not functional molecular markers directly related to target traits / genes (only linkage markers), resulting in invalid selection of target genes, or other non-target disadvantages caused by linkage redundancy genetic fragments etc.

Method used

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  • SNP locus combination for detecting resistance of tomato gray leaf spot disease and application thereof
  • SNP locus combination for detecting resistance of tomato gray leaf spot disease and application thereof
  • SNP locus combination for detecting resistance of tomato gray leaf spot disease and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0084] The screening of embodiment 1 SNP site combination

[0085] 1. Experimental materials

[0086] The variant group data of 660 representative tomato germplasm resources from different sources worldwide were selected for SNP site screening in this embodiment. The tomato variant group data is mainly based on the previous work of the project team where the inventor works ( Lin,T.,Zhu,G.,Zhang,J.et al.Genomic analyzes provide insights into the history of tomato breeding[J].Nat Genet,2014,46:1220~1226; Tieman D,Zhu G,Resende M F R, et al. A chemical genetic roadmap to improved tomato flavor [J]. Science, 2017, 355(6323): 391). Among them, the genotype data of some tomato germplasm resources and the resistance phenotype data of the Sm gene come from the existing public database (https: / / solgenomics.net / ).

[0087] 2. Screening of SNP site combinations

[0088] Use the target gene Sm (Solyc11g018710, Solyc11g018720 and Solyc11g018730, database source: https: / / solgenomics.net / ...

Embodiment 2

[0092] Example 2 Primer Synthesis and Kit Preparation

[0093] 1. Primer design and screening

[0094] According to the flanking sequences of the SNP01 site to SNP11 site provided in Example 1, for each SNP site, according to the KASP marker design and development principle (target product 80-150bp, primers that specifically match the target region on the reference genome and are located in non- SNP-intensive regions, avoid complex sequence regions with high A\T or G\C content, etc.), use Primer3.0 software to design two forward primers upstream of the SNP site, and design a reverse primer downstream.

[0095] Tomato samples to be tested include 21 randomly selected germplasm resources and breeding materials with known genotypes and phenotypes as resistant, susceptible or heterozygous controls, and finally add 3 ddH 2 O is used as the NTC blank control, a total of 24 copies.

[0096] DNA extraction Genomic DNA was extracted from the leaves of tomato samples by conventional C...

Embodiment 3

[0111] Example 3 Verification and Breeding Application of Tomato Gray Leaf Spot Resistance Gene Sm High Efficiency KASP Marker

[0112] In this example, based on the SNP loci provided in Example 1, the kit (including primer combinations) provided in Example 2 was used to carry out the verification and breeding application of the tomato gray leaf spot resistance gene Sm high-efficiency KASP marker.

[0113] Select 381 representative tomato samples including mainstream commercial varieties, core public germplasm of the National Resource Bank, breeding intermediate materials, new hybrid combinations, etc., involving large fruit tomatoes, cherry tomatoes, cluster tomatoes, fresh tomatoes, processed tomatoes and Among cultivated tomatoes of different types and origins such as farm species (landraces), 162 of them have obtained Sm genotype data through third-party commercial institutions using closely linked InDel markers (including 15 with known ash leaf spot resistance phenotypes ...

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Abstract

The invention relates to the technical field of plant biology, and particularly relates to an SNP locus combination for detecting resistance of tomato gray leaf spot disease and application thereof. Based on this, the invention develops a primer combination, a kit and a detection method capable of quickly, intuitively and effectively identifying the genotype state of a target SNP locus. The SNP locus combination can realize quick, accurate and high-throughput detection of a haplotype of an Sm segment of a resistance gene of the tomato gray leaf spot disease, has the advantages of simple operation, low cost, automation, high throughput efficiency, stable marking, safety, no toxicity, harmlessness and the like, can quickly and accurately identify the resistance of the tomato gray leaf spot disease at a high throughput in a tomato seedling stage, reduces the workload of artificial inoculation identification and field transplantation, improves the breeding efficiency, reduces the breeding cost, accelerates the breeding process, and is very suitable for modern commercial breeding application and large-scale genetic improvement research.

Description

technical field [0001] The invention relates to the field of plant biotechnology, in particular to a combination of SNP sites for detecting tomato gray leaf spot resistance and an application thereof. Background technique [0002] Tomato is an important vegetable economic crop in the world, which has important production application and basic research value. With the gradual expansion of tomato planting area around the world, the impact of tomato diseases and insect pests is increasing day by day. Among them, gray leaf spot disease (GLSD) of tomato is a new and popular fungal disease in recent years, and its incidence is increasing year by year. The causative agent is a fungus of the genus Stemphylium solani and S. lycopersici, which is a semi-known fungus. The disease occurs worldwide, especially in warm and humid regions. In recent years, it has generally occurred in spring and summer in the south and north of my country, especially in the cultivation of protected areas...

Claims

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Application Information

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IPC IPC(8): C12Q1/6895C12Q1/6858C12N15/11
CPCC12Q1/6895C12Q1/6858C12Q2600/13C12Q2600/156C12Q2531/113C12Q2563/107
Inventor 黄三文吴坤练群张金喆吕亚清
Owner AGRI GENOME INST OF SHENZHEN CHINESE ACADEMY OF AGRI SCI
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