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All Journal Jurnal Bios Logos BERITA BIOLOGI Jurnal Bioteknologi & Biosains Indonesia (JBBI) Makara Journal of Science
KRISTIANTO NUGROHO, KRISTIANTO
Balai Besar Penelitian dan Pengembangan Bioteknologi dan Sumber Daya Genetik Pertanian, Jl. Tentara Pelajar 3A, Bogor 16111 Indonesia Telp. (0251) 8337975
Agriculture, Biological Sciences & Forestry Biochemistry, Genetics & Molecular Biology Immunology & microbiology Medicine & Pharmacology

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KARAKTERISASI KERAGAMAN GENETIK 27 GENOTIPE CABAI BERDASARKAN MARKA SSR (SIMPLE SEQUENCE REPEAT) Terryana, Rerenstradika Tizar; Nugroho, Kristianto; Rijzaani, Habib; Lestari, Puji
BERITA BIOLOGI Vol 17, No 2 (2018)
Publisher : Research Center for Biology-Indonesian Institute of Sciences

Show Abstract | Download Original | Original Source | Check in Google Scholar | DOI: 10.14203/beritabiologi.v17i2.3313

Abstract

Chili pepper (Capsicum annuum) is one of the high economical horticultural comodity in Indonesia and its genetic diversity contributes to the success of breeding programs. Simple sequence repeat (SSR) markers can be used to analyze genetic diversity among chili pepper genotypes. The aim of this research was to analyze the genetic diversity of twenty-seven genotypes of chili pepper by using 24 SSR markers. The collected data was analyzed using cluster analysis and principle coordinate analysis (PCoA). The result showed that high allele variation (4–17 alleles) was observed among chili pepper genotypes tested, with an average allele number and Polymorphism Information Content (PIC) value was 7.708 and 0.758 (0.598–0.920) respectively. All of SSR markers showed PIC value >0.5 which indicated that these markers were suitable for chili pepper diversity studies with a high differentiation and with the average value of genetic diversity was 0.78. The clustering and principle coordinate analysis showed that twenty-seven genotypes of chili pepper were divided into two groups (coefficient of similarity 0.74 in cluster analysis) indicating a high genetic variability among them. Genetic diversity analysis in this study will be useful as an initial basis of selection for appropriate parents with desired traits to assist the breeding program of chili pepper in Indonesia.
METODE EKSTRAKSI DNA TANAMAN TANPA PRESIPITASI ETANOL UNTUK KEGIATAN POLYMERASE CHAIN REACTION (PCR) Nugroho, Kristianto; Terryana, Rerenstradika Tizar; Reflinur, .; Lestari, Puji
Jurnal Bioteknologi & Biosains Indonesia (JBBI) Vol 6, No 1 (2019): June 2019
Publisher : Badan Pengkajian dan Penerapan Teknologi (BPPT)

Show Abstract | Download Original | Original Source | Check in Google Scholar | Full PDF (196.537 KB) | DOI: 10.29122/jbbi.v6i1.3082

Abstract

A Simplified Plant DNA Extraction Protocol without Ethanol Precipitation for Polymerase Chain Reaction (PCR) Activities ABSTRACTMolecular-based research in agriculture includes DNA extraction stage involving DNA precipitation using ethanol or isopropanol which tends to take a long time. The purpose of this study was to obtain a plant DNA extraction method for Polymerase Chain Reaction (PCR) activities without going through the ethanol precipitation stage. Five important agricultural commodity crops, namely rice, corn, soybeans, chilies, and shallots were extracted by DNA using the modified Doyle and Doyle method. After the extraction phase using chloroform and isoamil alcohol solvents, the supernatant obtained was not precipitated using ethanol but was directly diluted and used as a template in PCR activities using two pairs of Simple Sequence Repeat (SSR) markers. The results showed that all samples could be well amplified, and amplicon tape visualized in both 1% agarose gel and 6% polyacrylamide gel were clearly visible. This method could save time and material, and reduce the dependence on liquid nitrogen. But this method is still limited to PCR requirements only, and cannot be used for activities that require high quality and quantity of DNA such as Next Generation Sequencing (NGS), digestion, and hybridization.Keywords: DNA extraction, ethanol precipitation, liquid nitrogen, PCR, SSR,  ABSTRAKPenelitian berbasis molekuler pada bidang pertanian mencakup tahapan ekstraksi DNA yang melibatkan presipitasi DNA menggunakan etanol atau isopropanol yang cenderung memakan waktu lama. Tujuan penelitian ini adalah untuk memperoleh metode ekstraksi DNA tanaman untuk kegiatan Polymerase Chain Reaction (PCR) tanpa melalui tahapan presipitasi etanol. Lima tanaman komoditas pertanian penting yaitu padi, jagung, kedelai, cabai, dan bawang merah diekstraksi DNA-nya menggunakan metode Doyle and Doyle yang dimodifikasi. Setelah tahap ekstraksi menggunakan pelarut kloroform dan isoamil alkohol, supernatan yang terbentuk tidak dipresipistasi menggunakan etanol melainkan langsung diencerkan dan digunakan sebagai template dalam kegiatan PCR menggunakan dua pasang marka Simple Sequence Repeat (SSR). Hasil menunjukkan bahwa seluruh sampel dapat teramplifikasi dengan baik serta pita hasil amplikon yang tervisualisasi baik pada gel agarosa 1% maupun gel poliakrilamid 6% terlihat jelas. Metode ini dapat menghemat waktu dan bahan serta mengurangi ketergantungan pemakaian nitrogen cair. Tetapi metode ini masih terbatas hanya untuk kebutuhan PCR saja dan tidak dapat digunakan untuk kegiatan yang membutuhkan DNA dengan kualitas serta kuantitas tinggi seperti Next Generation Sequencing (NGS), digesti, maupun hibridisasi.Kata Kunci: ekstraksi DNA, nitrogen cair, PCR, presipitasi etanol, SSR
METODE EKSTRAKSI DNA TANAMAN TANPA PRESIPITASI ETANOL UNTUK KEGIATAN POLYMERASE CHAIN REACTION (PCR) Nugroho, Kristianto; Terryana, Rerenstradika Tizar; Reflinur, .; Lestari, Puji
Jurnal Bioteknologi & Biosains Indonesia (JBBI) Vol. 6 No. 1 (2019): June 2019
Publisher : Balai Bioteknologi, Badan Pengkajian dan Penerapan Teknologi (BPPT)

Show Abstract | Download Original | Original Source | Check in Google Scholar | Full PDF (196.537 KB) | DOI: 10.29122/jbbi.v6i1.3082

Abstract

A Simplified Plant DNA Extraction Protocol without Ethanol Precipitation for Polymerase Chain Reaction (PCR) Activities ABSTRACTMolecular-based research in agriculture includes DNA extraction stage involving DNA precipitation using ethanol or isopropanol which tends to take a long time. The purpose of this study was to obtain a plant DNA extraction method for Polymerase Chain Reaction (PCR) activities without going through the ethanol precipitation stage. Five important agricultural commodity crops, namely rice, corn, soybeans, chilies, and shallots were extracted by DNA using the modified Doyle and Doyle method. After the extraction phase using chloroform and isoamil alcohol solvents, the supernatant obtained was not precipitated using ethanol but was directly diluted and used as a template in PCR activities using two pairs of Simple Sequence Repeat (SSR) markers. The results showed that all samples could be well amplified, and amplicon tape visualized in both 1% agarose gel and 6% polyacrylamide gel were clearly visible. This method could save time and material, and reduce the dependence on liquid nitrogen. But this method is still limited to PCR requirements only, and cannot be used for activities that require high quality and quantity of DNA such as Next Generation Sequencing (NGS), digestion, and hybridization.Keywords: DNA extraction, ethanol precipitation, liquid nitrogen, PCR, SSR,  ABSTRAKPenelitian berbasis molekuler pada bidang pertanian mencakup tahapan ekstraksi DNA yang melibatkan presipitasi DNA menggunakan etanol atau isopropanol yang cenderung memakan waktu lama. Tujuan penelitian ini adalah untuk memperoleh metode ekstraksi DNA tanaman untuk kegiatan Polymerase Chain Reaction (PCR) tanpa melalui tahapan presipitasi etanol. Lima tanaman komoditas pertanian penting yaitu padi, jagung, kedelai, cabai, dan bawang merah diekstraksi DNA-nya menggunakan metode Doyle and Doyle yang dimodifikasi. Setelah tahap ekstraksi menggunakan pelarut kloroform dan isoamil alkohol, supernatan yang terbentuk tidak dipresipistasi menggunakan etanol melainkan langsung diencerkan dan digunakan sebagai template dalam kegiatan PCR menggunakan dua pasang marka Simple Sequence Repeat (SSR). Hasil menunjukkan bahwa seluruh sampel dapat teramplifikasi dengan baik serta pita hasil amplikon yang tervisualisasi baik pada gel agarosa 1% maupun gel poliakrilamid 6% terlihat jelas. Metode ini dapat menghemat waktu dan bahan serta mengurangi ketergantungan pemakaian nitrogen cair. Tetapi metode ini masih terbatas hanya untuk kebutuhan PCR saja dan tidak dapat digunakan untuk kegiatan yang membutuhkan DNA dengan kualitas serta kuantitas tinggi seperti Next Generation Sequencing (NGS), digesti, maupun hibridisasi.Kata Kunci: ekstraksi DNA, nitrogen cair, PCR, presipitasi etanol, SSR
Pemanfaatan Teknologi Droplet Digital PCR (ddPCR) dalam Kegiatan Analisis Molekuler Tanaman Nugroho, Kristianto; Widyajayantie, Dwi; Ishthifaiyyah, Sayyidah Afridatul; Apriliani, Elisa
JURNAL BIOS LOGOS Vol 11, No 1 (2021): JURNAL BIOS LOGOS
Publisher : Universitas Sam Ratulangi

Show Abstract | Download Original | Original Source | Check in Google Scholar | DOI: 10.35799/jbl.11.1.2021.31101

Abstract

(Article History: Received 23 October 2020; Revised 9 January 2021; Accepted 18 January 2021) ABSTRAKSelama beberapa dekade terakhir, teknik PCR memberikan manfaat yang begitu besar dalam kegiatan penelitian di bidang biologi molekuler. Digital droplet PCR (ddPCR) merupakan salah satu teknologi PCR terbaru yang diklaim memiliki keunggulan dibanding teknik qPCR. Prinsip kerja teknik ini yaitu membagi sampel menjadi molekul-molekul kecil yang dipisahkan oleh emulsi minyak, air, dan senyawa penstabil sehingga membentuk droplets. Teknik ini memiliki kelebihan mampu melakukan kuantifikasi absolut maupun relatif pada DNA dengan konsentrasi sangat rendah, tidak memerlukan kurva standar, serta tidak sensitif terhadap kehadiran senyawa inhibitor. Teknik ini telah diaplikasikan pada kegiatan analisis molekuler tanaman di antaranya kegiatan pengukuran konsentrasi DNA dengan sangat akurat, deteksi kehadiran patogen pada jaringan tanaman, dan estimasi jumlah salinan T-DNA pada proses transformasi genetik.Kata kunci: PCR; droplet digital PCR; DNA; biologi molekuler; alat deteksi ABSTRACTOver the past decades, PCR technique has provided enormous benefits in molecular biology research activities. Digital droplet PCR (ddPCR) is one of the latest PCR technologies that is claimed to have advantages over the qPCR technique. The working principle of this technique is to divide the sample into small molecules, which separated by emulsions of oil, water, and stabilizing compounds to form droplets. This technique has the advantage of being able to perform absolute and relative quantification with very low DNA concentrations, does not require a standard curve, and less sensitive to the presence of inhibitor compounds. This technique has been applied to a number of plant molecular analysis, such as for measuring DNA concentrations very accurately, detecting the presence of pathogens in plant tissue, and estimating the copy number of T-DNA in the genetic transformation process.Keywords: PCR; droplet digital PCR; DNA; molecular biology; diagnostic tool.
The Use of Molecular Markers to Analyze the Genetic Diversity of Indonesian Pepper (Capsicum spp.) Varieties Based on Anthracnose Resistance Nugroho, Kristianto; Terryana, Rerenstradika T.; Manzila, Ifa; Priyatno, Tri Puji; Lestari, Puji
Makara Journal of Science
Publisher : UI Scholars Hub

Show Abstract | Download Original | Original Source | Check in Google Scholar

Abstract

Anthracnose is an important disease affecting the pepper plant and can lead to significant decreases in harvest yield. In this study, the genetic diversity of Indonesian pepper varieties was analyzed based on anthracnose resistance using molecular markers. DNA collected from 15 pepper varieties belonging to two species—Capsicum annuum L. and C. frutescens L.—was amplified using 14 molecular markers. The fungal isolate Colletotrichum capsici was inoculated into ripe harvested pepper fruits to observe their resistance to anthracnose as indicated by lesion size. Phylogenetic analysis revealed that the 15 pepper varieties could be classified into two major clusters with a genetic similarity coefficient of 0.63, and the pepper varieties exhibited varying degrees of resistance to anthracnose based on lesion size. Using the molecular markers, we were able to differentiate the species of pepper varieties, but not their resistance to anthracnose. All markers used in this study were confirmed to be highly informative (PIC > 0.5), suggesting their potential use in genetic studies on peppers. The marker GPMS29 was found to be significantly associated (P < 0.05) with anthracnose resistance. This information about the genetic diversity of peppers—along with the molecular markers used in our study—could prove to be useful in the further development of breeding programs of pepper plants in terms of anthracnose resistance in Indonesia.
Co-Authors . REFLINUR Apriliani, Elisa Dwi Widyajayantie HABIB RIJZAANI Ifa Manzila Ishthifaiyyah, Sayyidah Afridatul PUJI LESTARI RERENSTRADIKA T. TERRYANA, RERENSTRADIKA T. Rerenstradika Tizar Terryana, Rerenstradika Tizar Tri Puji Priyatno