Article | . 2017 Vol. 35, Issue. 3
Molecular Identification of Zoysia japonica and Zoysia sinica (Zoysia Species) Based on ITS Sequence Analyses and CAPS

Faculty of Biotechnology, Jeju National University1
Subtropical Horticulture Research Institute, Jeju National University2
Subtropical/tropical Organism Gene Bank, Jeju National Universtiy3
Department of Clinical Pathology, Cheju Halla College4
Department of Agronomy, Chungnam National University5
Omicsis. Inc. BVC, Korea Research Institute of Bioscience & Biotechnology,6

2017.. 344:360


Zoysiagrasses are important turf plants used for school playgrounds, parks, golf courses, and sports fields. The two most popular zoysiagrass species are Zoysia japonica and Zoysia sinica . These are widely distributed across different growing zones and are morphologically distinguishable from each other; however, it is phenotypically difficult to differentiate those that grow along the coastal line from those in beach area habitats. A combination of morphological and molecular approaches is desirable to efficiently identify these two plant cultivars. In this study, we used a rapid identification system based on DNA barcoding of the nrDNA-internal transcribed spacer (ITS) regions. The nrDNA-ITS regions of ITS1, 5.8S nrDNA, and ITS2 from Z. japonica , Z. sinica , Agrostis stolonifera , and Poa pratensis were DNA barcoded to classify these grasses according to their molecular identities. The nrDNA-ITS sequences of these species were found at 686 bp, 687 bp, 683 bp, and 681 bp, respectively. The size of ITS1 ranged from 248 to 249 bp, while ITS2 ranged from 270 to 274 bp. The 5.8S coding region ranged from 163 - 164bp. Between Z. japonica and Z. sinica , nineteen (2.8%) nucleotide sites were variable, and the G+C content of the ITS region ranged from 55.4 to 63.3%. Substitutions and insert/deletion (indel) sites in the nrDNA-ITS sequence of Z. japonica and Z. sinica were converted to cleaved amplified polymorphic sequence (CAPS) markers, and applied to the Zoysia grasses sampled to verify the presence of these markers. Among the 62 control and collected grass samples, we classified three groups: 36 Z. japonica , 22 Z. sinica , and 4 Z. japonica /Z. sinica hybrids. Morphological classification revealed only two groups; Z. japonica and Z. sinica . Our results suggest that used of the nrDNA-ITS barcode region and CAPS markers can be used to distinguish between Z. japonica and Z. sinica at the species level.

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