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Background <p>The molecular identification of maize varieties is important for protecting the legitimate rights and interests of farmers and maintaining national food security. However, scientific, simple, rapid, and systematic methods for germplasm identification remain limited, despite the critical need for accurate maize variety identification to prevent seed-market infringement and support agricultural security.</p> Results <p>In this study, we developed a high-throughput DNA barcoding system based on fluorescent simple sequence repeat (SSR) markers combined with capillary electrophoresis to distinguish 104 maize germplasms. After rigorous screening of 70 SSR primer pairs, 18 polymorphic loci distributed across eight chromosomes (Chr. 1–3, 5, and 7–10) were selected, demonstrating high discriminatory power, with an average polymorphism information content (PIC) value of 0.5791. The system showed strong sensitivity, with a detection limit of 0.125&#xa0;ng DNA; assay specificity, with no cross-reactivity with wheat, rice, or human DNA; and reproducibility across two platforms, GA118-16B and SeqStudio™ 8 Flex. Genetic diversity analysis identified 103 allelic variants, with observed heterozygosity ranging from 0.1635 to 0.8462. Unweighted pair-group method with arithmetic means (UPGMA) clustering and principal component analysis (PCA) grouped the materials into three categories that corresponded broadly to maturity period and ecological zone. A digital DNA molecular identity card was constructed by encoding SSR banding patterns into unique quick response (QR) codes, providing a practical tool for maize varietal authentication. In addition, an optimized multiplex polymerase chain reaction (PCR) protocol incorporating allele ladder standards was developed to ensure consistent genotyping under different experimental conditions.</p> Conclusion <p>This standardized SSR-based protocol provides a rapid, reproducible molecular identification approach for maize germplasm authentication and seed quality control, with potential application in variety protection and regulatory practice.</p>

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Construction of molecular identities and development of a strain identification kit based on fluorescent simple sequence repeat markers in maize

  • 期刊论文

  • 出版模式：

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  • 发表日期：

    2026年7月29日

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  • Shuozheng Mei,
  • Feng Wang,
  • Tao Zhang,
  • Zhiyong Fan,
  • Ailin Wan,
  • Jing Wang,
  • Ruihan Zhang,
  • Jiatong Xie,
  • Jiwei Xie,
  • Qingyou Xia,
  • Haibo Rong,
  • Genhong Wang

摘要

Background

The molecular identification of maize varieties is important for protecting the legitimate rights and interests of farmers and maintaining national food security. However, scientific, simple, rapid, and systematic methods for germplasm identification remain limited, despite the critical need for accurate maize variety identification to prevent seed-market infringement and support agricultural security.

Results

In this study, we developed a high-throughput DNA barcoding system based on fluorescent simple sequence repeat (SSR) markers combined with capillary electrophoresis to distinguish 104 maize germplasms. After rigorous screening of 70 SSR primer pairs, 18 polymorphic loci distributed across eight chromosomes (Chr. 1–3, 5, and 7–10) were selected, demonstrating high discriminatory power, with an average polymorphism information content (PIC) value of 0.5791. The system showed strong sensitivity, with a detection limit of 0.125 ng DNA; assay specificity, with no cross-reactivity with wheat, rice, or human DNA; and reproducibility across two platforms, GA118-16B and SeqStudio™ 8 Flex. Genetic diversity analysis identified 103 allelic variants, with observed heterozygosity ranging from 0.1635 to 0.8462. Unweighted pair-group method with arithmetic means (UPGMA) clustering and principal component analysis (PCA) grouped the materials into three categories that corresponded broadly to maturity period and ecological zone. A digital DNA molecular identity card was constructed by encoding SSR banding patterns into unique quick response (QR) codes, providing a practical tool for maize varietal authentication. In addition, an optimized multiplex polymerase chain reaction (PCR) protocol incorporating allele ladder standards was developed to ensure consistent genotyping under different experimental conditions.

Conclusion

This standardized SSR-based protocol provides a rapid, reproducible molecular identification approach for maize germplasm authentication and seed quality control, with potential application in variety protection and regulatory practice.