Isolation and genomic amplification of fish and shrimps from mangrove ecosystems in North Sumatra

Mangrove is a very typical ecosystem with a high level of diversity of flora and fauna. However, mangrove ecosystems are also one of the most vulnerable locations to extinction due to the high disturbances that occur. Some of the species that are susceptible to such disturbances are fish and shrimp. The use of barcode DNA is considered a fairly effective way to evaluate the condition of fish and shrimp populations in North Sumatra. Therefore, this study aims to obtain information on the success of kit methods and amplification patterns of barcoding marks 16S for fish and shrimp in mangrove ecosystems North Sumatra. The results showed that the kit extraction method produced a pattern of band that varied thick, thin and even smear. PCR amplification results with the 16S gene showed a good band pattern and indicated that the 16S primer used can detect fish and shrimp species that are visualized in the form of DNA bands.

The Identification of Several Dipterocarpaceae and Fagaceae Trees by Barcode DNA Coupled with High-Resolution Melting Analysis

The loss of forests is a major environmental, social, and economic problem. The disappearance has been occurring to an extreme degree in many parts of Southeast Asia, including Thailand. Logging and clearing of forests for agriculture, cash crops, and food production has destroyed much of the tropical forests in Thailand. Floristic inventory could provide essential information for forest conservation but species identification as a part of inventory creating could be challenging in some cases. Barcode DNA coupled with High Resolution Melting analysis (Bar-HRM) was used here in aiding species identification of plant in Dipterocarpaceae (Dipterocarpus alatus, D. costatu, D. intricatus, D. obtusifolius, Hopea ferrea, H. odorata, Shorea guiso, S. obtuse, S. roxburghii, and S. siamensis) and Fagaceae (Castanopsis echinocarpa, C. inermis, Lithocarpus wallichianus, Quercus aliena and Q. oidocarpa) families. Two main experiments were conducted including: (1) a comparing method for primer design and (2) testing the robustness of the Bar-HRM by trying to identify tree samples that did not have sequences in the GenBank. In experiment 1, the manual design primer pair was found to be the best fit for the work. Of key importance is finding the primers which give the most nucleotide variations within the generated amplicon; this is a parameter that cannot be set in any web-based tools. Next, in experiment 2, Bar-HRM using primers of ITS1 and ITS2 regions were able to discriminate all 10 tested tree species without any problem, even when there were no sequences of the samples to be analysed before performing the HRM. Here, Bar-HRM poses potential to be a game-changer in tropical forest conservation, as it will be useful for species identification.

DNA Barcoding Dalugha (Cyrtosperma Merkusii) di Kepulauan Talaud dan Minahasa Selatan Berdasarkan Gen rbcL

(Article History: Received June 28, 2021; Revised August 30, 2021; Accepted Sept 3, 2021) ABSTRAKDalugha (Cyrtospera merkusii (Hassk.)Schott) merupakan tanaman endemik Sulawesi Utara yang digunakan sebagai pangan alternatif (penganti beras). Penelitian ini bertujuan untuk membandingkan spesies daluga di Kepulauan Talaud dan Minahasa Selatan menggunakan DNA barcode gen rbcL (ribulose 1,5 bisphosphate carboxylase large). Perbandingan barcode DNA yang dilakukan pada empat sampel yang berbeda lokasi tersebut keduanya menghasilkan tingkat kesamaan 100% (identik). Dengan demikian, tidak ada variasi intra spesies yang ditemukan dari semua sampel yang ada. Selanjutnya, kemiripan sampel-sampel ini telusuri kemiripannya dengan kerabat terdekat yang tercatat di GenBank menggunakan BLAST (Basic Local Alignment Search Tool). Tanaman dalugha dalam penelitian ini memiliki kemiripan 99,82% dengan tumbuhan Anaphyllopsis americana (AM905753.1), dan kemiripannya 99,63% dengan Cyrtosperma macrotum (AM905750.1), Lasimorpha senegalensis (AM905755.1), Pycnospatha arietina (AM905751.1), dan Podolasia stipitata (AM905752.1). Belum ada rekor sekuens DNA gen rbcL dari spesies ini yang dibisa dibandingkan di GenBank.Kata Kunci: Dalugha; DNA barcoding; gen rbcL ABSTRACTDalugha (Cyrtospera merkusii (Hassk.) Schott) is an endemic plant in North Sulawesi that is used as alternative food (substitute for rice). This research aimed to compare the DNA barcode of dalugha in Talaud Islands and in South Minahasa using rbcL (ribulose 1,5 bisphosphate carboxylase large) gene. The DNA barcoding comparison of all four samples in both area resulted in 100% similarity (identical). Therefore, there is no intraspecific variation found in all samples. Furthermore, the similarity of these samples were conducted with BLAST (Basic Local Alignment Search Tool) to compare with its closest relatives in GenBank. The closest relatives of this plant, based on similarity information, are 99.82% with Anaphyllopsis americana (AM905753.1) and all 99.63% with Cyrtosperma macrotum (AM905750.1), Lasimorpha senegalensis (AM905755.1), Pycnospatha arietina (AM905751.1), and Podolasia stipitata (AM905752.1). There is no record yet of rbcL gene sequence of C. merkusii in GenBank for comparison.Keywords: Dalugha; DNA barcoding; rbcL gene

Deteksi dan Identifikasi Nematoda Puru Akar (Meloidogyne spp.) pada Tanaman Bit Menggunakan Metode DNA Barcoding

Identifikasi nematoda berdasarkan karakter morfologi memerluan ketelitian tinggi dan perlu didukung hasil identifikasi secara molekuler. Salah satu metode identifikasi molekuler ialah dengan DNA barcode. DNA barcode merupakan identifikasi molekuler dengan menggunakan gen cytochrome oksidase sub unit 1 (CO1). Penelitian ini bertujuan untuk mengidentifikasi NPA yang berasosiasi pada tanaman bit berdasarkan PCR dan perunutan nukleotida gen CO1 Meloidogyne. Sampel umbi bit diperoleh dari lahan pertanian yang terletak di Kecamatan Cipanas, Kabupaten Cianjur, Jawa Barat. Pengambilan sampel dilakukan dengan metode purposive sampling. Identifikasi morfologi dengan mengamati pola perineal NPA betina. Identifikasi molekuler meliputi ekstraksi, amplifikasi, visualisasi DNA, analisis peruntutan nukleotida dan filogenetika. Hasil pengamatan pola perineal secara morfologi didapatkan spesies M. incognita dan M. arenaria. Amplifikasi DNA yang dilakukan menggunakan primer spesifik CO1SIF (5’-GCCTGCATTTGGTTAG-‘3) dan CO1SIR (5’-TCAAACCAGTCCT-‘3), CO1SAF (5’-GGGTACTGGATGAACATTA-‘3) dan CO1SAR (5’-ACTTCAGGATGACCAAA-‘3)berhasil mendapatkan pita DNA berukuran ± 360 untuk M. arenaria dan ± 326 untuk M. incognita. Hasil sekuensing menunjukkan bahwa isolat M. incognita asal Indonesia berkerabat dekat dengan isolat M. incognita asal Cina, Amerika Serikat, Vietnam, dan Inggris dengan tingkat homologi 100%. Hasil analisis filogenetika menunjukkan bahwa M. incognita Indonesia masih satu kelompok dengan M. incognita asal Cina, Amerika Serikat, Vietnam dan Inggris. Adapun isolat M. arenaria berkerabat dekat dengan isolat M. arenaria asal Argentina dan Amerika Serikat dengan tingkat homologi 100%. Hasil analisis filogenetika menunjukkan bahwa M. arenaria asal Indonesia masih satu kelompok dengan M. arenaria asal Argentina dan Amerika Serikat.

The DNA barcode of red fruit pandan (Pandanaceae) cultivar from Wamena, Papua Province, Indonesia based on matK gene

Zebua LI, Gunaedi T, Budi IM, Lunga N. 2019. The DNA barcode of red fruit pandan (Pandanaceae) cultivar from Wamena, Papua Province, Indonesia based on matK gene. Biodiversitas 20: 3405-3412. The red fruit pandan has been used by the people of Wamena in central highlands of Papua as medicinal plants, food ingredients, and religious ceremonies. Based on morphological characters, there are 39 cultivars of red fruit pandan in New Guinea. A standard method for plant species identification through DNA barcodes has been recommended to use matK gene. The research aimed to determine similarities in DNA barcode sequences of six red fruit pandan cultivars with its close relative that listed in NCBI system and to recommend the reliable DNA barcode for identification of this cultivar. Polymerase Chain Reaction was employed DNA to amplify matK gene fragments using an available forward primer (5’CGA TCT ATT CAT TCA ATA TTT C 3’) and reverse primer (5’TCT AGC ACA CGA AAGTCG AAG T 3’). The software BLAST, Bioedit, and ClustalX were used to analyze the data. Barcode DNA of red fruit pandan showed 1474 bp nucleotides sequence based on matK gene. An indication of six red fruit pandan cultivars showed that high similarity 99% with Pandanus conoideus Lam. It can be concluded that matK gene can be used to determine the species level in Pandanus.

Identifying accurate metagenome and amplicon software via a meta-analysis of sequence to taxonomy benchmarking studies

Metagenomic and meta-barcode DNA sequencing has rapidly become a widely-used technique for investigating a range of questions, particularly related to health and environmental monitoring. There has also been a proliferation of bioinformatic tools for analysing metagenomic and amplicon datasets, which makes selecting adequate tools a significant challenge. A number of benchmark studies have been undertaken; however, these can present conflicting results. In order to address this issue we have applied a robustZ-score ranking procedure and a network meta-analysis method to identify software tools that are consistently accurate for mapping DNA sequences to taxonomic hierarchies. Based upon these results we have identified some tools and computational strategies that produce robust predictions.

Status and challenges for the molecular characterization of Fannia (Insecta, Diptera, Fanniidae) in Brazil

Both the advancement of technology associated with molecular characterization studies of species have contributed to the increased use of the Barcode DNA method for identifying individuals. For some areas, such as forensics, species determination is essential for information regarding the time of death. In this study, Fannia nucleotide sequences available in public databases were quantified and used to evaluate the scenario and the challenge to establish molecular research with fanids in Brazil.