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AbstrakDendrobium linearifolium Teijsm. & Binn. merupakan salah satu spesies anggrek yang telah dimanfaatkan sebagai obat herbal tradisional di Bali. Bagian pseudobulb dari D. linearifolium dimanfaatkan oleh masyarakat Bali untuk mengobati sakit telinga. Jenis anggrek ini menunjukkan kemiripan morfologi dengan anggrek Dendrobium faciferum J.J. Sm. dan Dendrobium crumenatum Sw. sehingga menyulitkan dalam proses identifikasi menggunakan karakter morfologi. Penggunaan DNA barcoding dengan penanda molekuler Internal Transcribed Spacer 2 (ITS2) diharapkan dapat digunakan untuk membedakan D. linearifolium secara akurat dan tepat. Penelitian ini bertujuan untuk mengidentifikasi D. linearifolium menggunakan DNA barcoding ITS2 sebagai penanda molekuler DNA inti. DNA genom D. linearifolium Teijsm. & Binn. diisolasi dan digunakan sebagai DNA template dalam reaksi PCR. Amplikon yang dihasilkan kemudian diurutkan dengan cara sekuensing. Hasil penelitian menunjukkan urutan sekuen ITS2 dari D. linearifolium Teijsm. & Binn. memiliki homologi tinggi  dengan D. junceum (Accession: AB593590; Per. Ident: 92,57%). Hasil analisis bioinformatika menunjukkan urutan sekuen ITS memiliki variasi basa nitrogen yang tinggi dan spesifik yang menunjukkan ciri khas suatu spesies. Dengan demikian, urutan ini dapat dijadikan sebagai penanda molekuler dalam identifikasi spesies anggrek. Sebagai kesimpulan, sekuen ITS2 dapat direkomendasikan sebagai penanda molekuler untuk identifikasi anggrek D. linearifolium Teijsm. & Binn.Abstract Dendrobium linearifolium Teijsm. & Binn is a species of orchid that has been used as traditional herbal medicine in Bali. Balinese uses the pseudobulb of D. linearifolium to treat earaches. This type of orchid shows morphological similarities to D. faciferum J.J. Sm. and D. crumenatum Sw. which raised ardurous to identify using morphological characters. DNA barcoding with the Internal Transcribed Spacer 2 (ITS2) as molecular marker is expected to distinguish D. linearifolium accurately and precisely. This study aims to identify D. linearifolium using ITS2 as a molecular marker. The Genomic DNA of D. linearifolium was extracted and used as template DNA in PCR reactions. DNA amplicons are then sequenced. The results showed that ITS2 sequence of D. linearifolium Teijsm. & Binn. has high homology with D. junceum (Accession: AB593590; Per. Ident: 92.57%). The results of the bioinformatics analysis showed that the ITS sequence had a high variation and specificity of nitrogen base to indicate the characteristics of a species. Therefore, this sequence can be used as a molecular marker to identify an orchid to species. In conclusion, the ITS2 sequence can be recommended as a molecular marker for species identification of D. linearifolium.

Indonesia has high biodiversity for plant species, including orchids with medicinal potential such as Phalaenopsis deliciosa. Generally, morphological characters, especially in flowers are used for orchids identification. However, when the plants are not in the flowering period, the identification becomes difficult. Therefore an alternative method, such as molecular identification (DNA barcoding) needs to be applied for the best solution. This research, which was conducted with three different markers found that the identity level of matK, rbcL, and ITS to other orchids species was 99-98%, 98%, and 94-96%, respectively. Furthermore, matK and ITS showed high specificity for Phalaenopsis deliciosa, and are therefore recommended as the best molecular identification marker of genus Phalaenopsis.

Molecular identification using short orthological DNA sequences (DNA barcoding) has been applied for the classification of orchid species which is a major step in biodiversity management, conservation, breeding, authenticating components of herbal products, and tracking the adulteration of orchid species. One of the most widely used loci for phylogenetic inference at the generic and infrageneric levels in plants was ITS located between 18S rDNA, 5.8S rDNA, and 26S rDNA. The ITS or ITS2 region has been suggested as a plant barcode in some previous studies. However, DNA barcoding using the entire ITS genome is considered less effective and efficient in the process of PCR amplification and sequencing. Besides, complete DNA barcodes are difficult to obtain from herbarium samples and herbal products because some DNA sequences have been degraded. DNA mini-barcodes were developed over the past ten years to overcome issues related to DNA barcoding. DNA mini-barcodes use shorter DNA segments for PCR amplification, so they can identify species effectively and efficiently compared to the regular DNA barcoding. Specific primers that encode the ITS1 and ITS2 regions need to be designed for the PCR amplification. DNA mini-barcoding ITS has proven useful in species identification, classification studies, and authentication of specific orchid species. Therefore, a new rapid identification method based on the ITS mini-barcode is expected to be established, especially for orchid species. Keywords : DNA mini-barcode, Internal Transcribed Spacer, Orchid, Primer Design