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Molecules 2017, 22(4), 665; doi:10.3390/molecules22040665

Development of User-Friendly Method to Distinguish Subspecies of the Korean Medicinal Herb Perilla frutescens Using Multiplex-PCR

1
Jeonnam Institute of Natural Resources Research, Jangheung-gun, Jeollanamdo 59338, Korea
2
School of Biological Sciences and Technology, Chonnam National University, Gwangju 61186, Korea
3
Department of Pharmacy, College of Pharmacy, Mokpo National University, Muan, Jeonnam 58554, Korea
*
Author to whom correspondence should be addressed.
Academic Editor: Derek J. McPhee
Received: 8 March 2017 / Revised: 8 April 2017 / Accepted: 19 April 2017 / Published: 21 April 2017
(This article belongs to the Section Molecular Diversity)
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Abstract

Perilla (Perilla frutescens) is an economically and culturally important plant in East Asia. Plant breeding between cultivars has enhanced the genetic diversity of perilla overall, but means that functionally diverse subspecies are more difficult to identify and distinguish. In this study, we developed gene-based DNA markers to distinguish between the Korean herbal medicinal perilla varieties. We identified informative simple sequence repeat (SSR) regions on the promoter regions of the Myb-P1 and dihydroflavonol 4-reductase (DFR) genes, as well as a large insertion-deletion (indel) region in the limonene synthase (LS) gene, and developed markers to characterize the distinct subspecies differences (PfMyb-P1pro, PfDFRpro, and PfLS, respectively). Using the PfLS primers, a 430-bp region could be amplified from P. frutescens var. acuta, crispa, and f. viridis (known as Jasoyeop, Jureum-soyeop, and Chungsoyeop, respectively), but not from P. frutescens var. japonica (Dlggae). The PfMybpro primers resulted in PCR products of 314 or 316, 330, 322, and 315 bp from Dlggae, Jasoyeop, Jureum-soyeop, and Chungsoyeop, respectively, and the PfDFRpro primers resulted in products of 189 or 202, 187 or 189, 185 or 189, and 193bp, respectively, for the four perilla subspecies. Combining these three reactions into a single multiplex PCR approach resulted in subspecies-specific PCR band patterns for six common types of commercial perilla, distinguishing between three varieties of Dlggae (Cham-Dlggae, Ip-Dlggae, and Bora-Dlggae), as well as identifying Jasoyeop, Jureum-soyeop, and Chungsoyeop. These user-friendly markers will be valuable as a simple and efficient method for identifying the Korean medicinal herb Jasoyeop, as well as distinguishing between other functionally distinct subspecies, which may have broad applications in the Korean herbal industry. View Full-Text
Keywords: simple sequence repeat (SSR); Myb-P1; dihydroflavonol 4-reductase (DFR); Perilla; Jasoyeop; indel marker simple sequence repeat (SSR); Myb-P1; dihydroflavonol 4-reductase (DFR); Perilla; Jasoyeop; indel marker
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This is an open access article distributed under the Creative Commons Attribution License which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. (CC BY 4.0).

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Kim, Y.; Kim, A.-Y.; Jo, A.; Choi, H.; Cho, S.-S.; Choi, C. Development of User-Friendly Method to Distinguish Subspecies of the Korean Medicinal Herb Perilla frutescens Using Multiplex-PCR. Molecules 2017, 22, 665.

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