국내 작약 품종을 저비용으로 식별하기 위한 InDel 마커 개발
Development of InDel Markers for the Identification of Herbaceous Peony Varieties
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Background: The herbaceous peony (Paeonia lactiflora) is widely used as an ornamental and medicinal crop in Korea; however, morphology-based cultivar discrimination is unreliable, high- lighting the need for low-cost markers to facilitate the rapid identification, maintenance, distribu- tion, and quality control of cultivars. Methods and Results: Next-generation sequencing was performed for five Korean cultivars (Uis- eong, Taebaek, Sagok, Daecheong, and Geopung). Putative insertion/deletion (InDel) sites, mostly ≥19 bp, were detected by comparative analysis using CLC Genomics Workbench software, and primers were designed for 22 loci. Of these, nine markers yielded clear amplicon length polymor- phisms that were resolved using simple PCR–agarose gels. In total, 18 distinct three-marker com- binations uniquely identified all five cultivars; for example, PLID1-5/PLID5-2/PLID8-2. Weak upper bands at some loci were confirmed as heteroduplexes by T7 endonuclease I and were excluded from scoring. The Fragment Analyzer profiles were fully concordant with the gel results. Conclusion: These InDel markers enable reliable, low-cost, and gel-based identification of Korean herbaceous peony cultivars, and are applicable for variety protection, purity testing, and distribu- tion authentication.
Abstract
Background: The herbaceous peony (Paeonia lactiflora) is widely used as an ornamental and medicinal crop in Korea; however, morphology-based cultivar discrimination is unreliable, high- lighting the need for low-cost markers to facilitate the rapid identification, maintenance, distribu- tion, and quality control of cultivars. Methods and Results: Next-generation sequencing was performed for five Korean cultivars (Uis- eong, Taebaek, Sagok, Daecheong, and Geopung). Putative insertion/deletion (InDel) sites, mostly ≥19 bp, were detected by comparative analysis using CLC Genomics Workbench software, and primers were designed for 22 loci. Of these, nine markers yielded clear amplicon length polymor- phisms that were resolved using simple PCR–agarose gels. In total, 18 distinct three-marker com- binations uniquely identified all five cultivars; for example, PLID1-5/PLID5-2/PLID8-2. Weak upper bands at some loci were confirmed as heteroduplexes by T7 endonuclease I and were excluded from scoring. The Fragment Analyzer profiles were fully concordant with the gel results. Conclusion: These InDel markers enable reliable, low-cost, and gel-based identification of Korean herbaceous peony cultivars, and are applicable for variety protection, purity testing, and distribu- tion authentication.