Combination of SNP loci for detection of tomato yellow leaf curl virus disease resistance and its application

A technology for tomato yellowing and curving leaves and viral diseases, applied in the field of plant biology, can solve the problems of linkage burden, ineffective selection of target genes, and insufficient versatility of SNP sites, etc., achieve accurate detection, improve breeding efficiency, and reduce manual inoculation identification and field transplanting workload

Active Publication Date: 2022-06-24
AGRI GENOMICS INST 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 upstream and downstream adjacent regions of the known Ty-1 / Ty-3 and Ty-2 genes and within the genes, multiple genes that can quickly and effectively detect the resistance to tomato yellow leaf curl virus disease were obtained. SNP loci can realize accurate identification and selection of target gene regions, thereby solving the lack of versatility of SNP loci in the prior art, or functional molecular markers that are not directly related to target traits / genes (only linkage markers), causing target gene Selection is invalid, or linkage drag brings other non-target unfavorable gene fragments and other problems

Method used

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  • Combination of SNP loci for detection of tomato yellow leaf curl virus disease resistance and its application
  • Combination of SNP loci for detection of tomato yellow leaf curl virus disease resistance and its application
  • Combination of SNP loci for detection of tomato yellow leaf curl virus disease resistance and its application

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0080] Example 1 Screening of SNP site combinations

[0081] 1. Experimental materials

[0082] The variation group data of 660 representative tomato germplasm resources worldwide, from different source types, are selected for the SNP site screening in this example. The source of the tomato variation group data is mainly based on the preliminary work of the project team where the inventor is located. (Lin, T., Zhu, G., Zhang, J. et al. Genomic analyses 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 Ty-1 / Ty-3 and Ty-2 genes were obtained from the existing public database (https: / / solgenomics.net / ).

[0083] 2. Screening of SNP site combinations

[0084] Using target genes Ty-1 / Ty-3 genes (Solyc06g051170, Sol...

Embodiment 2

[0088] Example 2 Primer synthesis and kit preparation

[0089] 1. Primer design and screening

[0090] According to the flanking sequences of Ty1-SNP01 site~Ty1-SNP08 site and Ty2-SNP01 site~Ty2-SNP08 site provided in Example 1, for each SNP site, adopt the design and development principle according to KASP marker (target product 80-150bp, the primers specifically match the target region on the reference genome and are located in non-SNP-intensive regions, avoiding complex sequence regions with high A\T or G\C content, etc.), using Primer3.0 software to design two upstream of the SNP site Forward primer, downstream design a reverse primer.

[0091] The tomato samples to be tested include some germplasm resources with known genotypes and yellow leaf curl virus disease resistance phenotypes and 21 randomly selected samples from the breeding materials as resistance, sensitivity or heterozygous controls, and finally add 3 ddH 2 O as the NTC blank control, a total of 24 copies. ...

Embodiment 3

[0109] Example 3 Validation and breeding application of highly efficient KASP markers for tomato yellow leaf curl virus disease resistance genes Ty-1 / Ty-3 and Ty-2

[0110] Based on the SNP loci provided in Example 1, this example uses the kit (including primer combination) provided in Example 2 to carry out high-efficiency KASP of tomato anti-yellowing leaf curl virus disease genes Ty-1 / Ty-3 and Ty-2 Validation of markers works with breeding applications.

[0111] Selected 381 representative tomato samples including mainstream commercial varieties, core public germplasm of the national resource bank, intermediate breeding materials, new hybrid combinations, etc., involving large fruit tomatoes, cherry tomatoes, bunch tomatoes, fresh tomatoes, processed tomatoes and Farming varieties (landraces) and other types of cultivated tomatoes, 154 of which have obtained Ty-1 / Ty-3 and Ty-2 genotype data through third-party commercial institutions using closely linked SCAR markers (inclu...

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Abstract

The invention relates to the field of plant biotechnology, in particular to a combination of SNP sites for detecting tomato yellow leaf curl virus disease resistance and an application thereof. Based on this, the present invention develops a primer combination, a kit and a detection method that can quickly, intuitively and effectively identify the genotype status of the target SNP site. Application of the present invention can realize rapid, accurate and high-throughput detection of haplotypes of tomato yellow leaf curl virus disease resistance genes Ty-1 / Ty-3 and Ty-2, and has the advantages of simple operation, low cost, and With the advantages of automation, high flux efficiency, stable labeling, safety, non-toxicity and harmlessness, etc., it can quickly, accurately and high-throughput identify the resistance of tomato yellow leaf curl virus disease at the tomato seedling stage, reducing manual inoculation identification and field transplantation. It reduces the workload of breeding, improves breeding efficiency, reduces breeding costs, and accelerates the breeding process. It is very suitable for modern commercial breeding applications 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 resistance to yellow leaf curl virus disease of tomato and its application. Background technique [0002] Tomato is an important vegetable economic crop in the world, with important production application and basic research value. With the gradual expansion of tomato planting area around the world, the impact of tomato pests and diseases is increasing day by day. Among them, tomato yellow leaf curl virus (TYLCV) is a virus disease of Solanaceae crops widely distributed in tropical and subtropical regions. It is mainly carried and spread by Bemisia tabaci. After the tomato is infected, the plants show obvious dwarfing, the leaf margins turn yellow, the leaves become smaller and curled, which seriously damages the growth, flowering and fruit setting. Exterminated. Tomato yellow leaf curl virus disease resistance is mainly controlled by ...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12Q1/6858C12Q1/6895C12N15/11
CPCC12Q1/6858C12Q1/6895C12Q2600/156C12Q2600/13C12Q2600/172C12Q2563/107C12Q2531/113
Inventor 黄三文吴坤张金喆练群胡勇
Owner AGRI GENOMICS INST CHINESE ACADEMY OF AGRI SCI
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