Scientia Agricultura Sinica ›› 2025, Vol. 58 ›› Issue (22): 4746-4756.doi: 10.3864/j.issn.0578-1752.2025.22.014

• HORTICULTURE • Previous Articles     Next Articles

Development and Application of A KASP Marker-Based Identification System for Tomato Varieties

SU XiaoMei(), YANG ZongHui, LIU ShuMei, ZHANG ZongJie, LÜ HongJun, HOU LiXia()   

  1. Institute of Vegetables, Shandong Academy of Agricultural Sciences/Key Laboratory of Huang Huai Protected Horticulture Engineering, Ministry of Agriculture and Rural Affairs/Shandong Key Laboratory of Bulk Open-Field Vegetable Breeding, Jinan 250100
  • Received:2025-05-26 Accepted:2025-07-14 Online:2025-11-16 Published:2025-11-21
  • Contact: HOU LiXia

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Abstract:

【Objective】To screen KASP core markers and establish a high-throughput molecular identification system for tomato, providing technical support for tomato variety authentication, purity testing, and plant variety protection. 【Method】Based on re-sequencing data of tomato inbred lines, high-quality SNP loci covering the whole genome were selected for KASP markers design. A total of 360 tomato varieties, collected from different breeding research institutions and private companies, were used to validate the designed KASP markers. According to genotyping data, a set of KASP core markers with high polymorphism information content (PIC), excellent stability, and relatively uniform distribution across chromosomes was screened. Cluster analysis and DNA fingerprinting of the collected tomato varieties were performed, and the core markers were applied to authenticate variety authenticity and assess seed purity. 【Result】Utilizing the SNPs obtained from re-sequencing data analysis of over 300 tomato materials, 600 SNP loci were selected for marker development, averaging 50 per chromosome. Among these, 517 were successfully converted into KASP markers, with a conversion success rate of 86.2%. Based on physical distances on chromosomes, 10-12 markers per chromosome were prioritized, totaling 128 markers, which were validated and refined using diverse tomato varieties. Ultimately, 68 core primer pairs were selected, averaging 4-8 per chromosome. Genotyping analysis of 360 tomato varieties using these 68 core markers revealed similarity coefficients ranging from 0.32 to 1.00 in cluster analysis. For four tomato types (fresh tomatoes, flavor tomatoes, cherry tomatoes and processing tomatoes), the core marker sets were streamlined to 48, 32, 36, and 36 markers, respectively, with PIC values all exceeding 0.35. These markers effectively distinguished different varieties. To validate marker applicability, two flavor tomato varieties were authenticated using core markers, revealing two differing loci, confirming their distinctness. Additionally, two polymorphic markers between parental lines were used to assess the purity of hybrid seeds of Pinfan 4031, yielding 100% purity, consistent with field evaluation results. 【Conclusion】The developed KASP markers are applicable for tomato genetic diversity analysis, DNA fingerprinting, variety authenticity testing, and hybrid seed purity evaluation.

Key words: tomato, KASP, core markers, fingerprint, variety identification

Fig. 1

The physical position of 68 KASP markers on the tomato reference genome (SL4.0) The red represents the centromere area"

Fig. 2

Representative results of KASP genotyping assay a: KASP marker that can be used, have good typing and a high PIC value; B: KASP marker that can be used in cherry tomatoes, have classification differences among different types of tomatoes (line A-E for cherry tomatoes and F-H for flavor tomatoes); c: KASP marker that cannot be used, have good typing but a low PIC value"

Fig. 3

The fingerprint of 360 tomato varieties based on 68 KASP markers B: Fresh tomato; K: Flavor tomato; P: Processing tomato; Y: Cherry tomato. The same as below. In the fingerprint map, different colors represent the homozygous type consistent with the reference genome (0, red), heterozygous type (1, light pink), variant homozygous type (2, blue), and unknown genotype (NA, gray), respectively"

Fig. 4

Cluster map of 360 tomato varieties based on UPGMA"

Fig. 5

The authenticity identification of two flavor tomato varieties based on a core set of 32 markers"

Fig. 6

Identification results of markers 3-12 (a) and 7-2 (b) on the parents and F1 hybrids of Pinfan 4031"

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