The groundnut breeding program at International Crops Research Institute for the Semi-Arid Tropics routinely performs marker-based early generation selection (MEGS) in thousands of segregating populations. The existing MEGS includes planting of segregating populations in fields or glasshouses, label tagging, and sample collection using leaf-punch from 20–25 day old plants followed by genotyping with 10 single nucleotide polymorphisms based early generation selection marker panels in a high throughput genotyping (HTPG) platform. The entire process is laborious, time consuming, and costly. Therefore, in order to save the time of the breeder and to reduce the cost during MEGS, we optimized a single seed chipping (SSC) process based MEGS protocol and deployed on large scale by genotyping >3000 samples from ongoing groundnut breeding program. In SSC-based MEGS, we used a small portion of cotyledon by slicing-off the posterior end of the single seed and transferred to the 96-deep well plate for DNA isolation and genotyping at HTPG platform. The chipped seeds were placed in 96-well seed-box in the same order of 96-well DNA sampling plate to enable tracking back to the selected individual seed. A high germination rate of 95–99% from the chipped seeds indicated that slicing of seeds from posterior end does not significantly affect germination percentage. In addition, we could successfully advance 3.5 generations in a year using a low-cost rapid generation turnover glass-house facility as compared to routine practice of two generations in field conditions. The integration of SSC based genotyping and rapid generation advancement (RGA) could significantly reduce the operational requirement of person-hours and expenses, and save a period of 6–8 months in groundnut genetics and breeding research.

中文翻译：

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用于加速花生遗传学和育种研究的基于单种子的高通量基因分型和快速世代推进

国际半干旱热带作物研究所的花生育种计划定期在数千个隔离种群中进行基于标记的早期世代选择 (MEGS)。现有的 MEGS 包括在田间或温室中种植隔离种群、标记标签和使用 20-25 天龄植物的叶片采集样本收集，然后在高通量基因分型中使用 10 个基于单核苷酸多态性的早期选择标记面板进行基因分型。 HTPG) 平台。整个过程费力、费时、成本高。因此，为了节省育种者的时间并降低 MEGS 期间的成本，我们优化了基于 MEGS 协议的单种子切片 (SSC) 过程，并通过对正在进行的花生育种计划的 > 3000 个样本进行基因分型进行了大规模部署。在基于 SSC 的 MEGS 中，我们通过切下单个种子的后端使用一小部分子叶，并将其转移到 96 深孔板中，用于在 HTPG 平台上进行 DNA 分离和基因分型。将切碎的种子按照与 96 孔 DNA 采样板相同的顺序放置在 96 孔种子盒中，以便能够追踪回所选的单个种子。切碎的种子的发芽率高达 95-99%，表明从后端切下种子不会显着影响发芽率。此外，与现场条件下两代的常规做法相比，我们可以使用低成本的快速发电周转温室设施在一年内成功推进 3.5 代。