scholarly journals Environmental DNA (eDNA) reveals endangered narrow sawfish across Indonesian Reefs

2021 ◽  
Vol 944 (1) ◽  
pp. 012020
Author(s):  
L M I Sani ◽  
A K Husna ◽  
B Subhan ◽  
H Madduppa
Keyword(s):  
Filter Paper ◽  
Pcr Amplification ◽  
Environmental Dna ◽  
Marine Species ◽  
Visual Census ◽  
Survey Tool ◽  
Polymerase Chain ◽  
Seawater Samples ◽  
Next Generation Sequencing Ngs ◽  
Generation Sequencing

Abstract Environmental DNA or eDNA is a powerful method to uncover marine organisms in the seawaters. Furthermore, many marine species are difficult to determine in the sea waters because of their rare existence based on the visual census. In this study, we implemented environmental DNA to investigate the presence of the endangered species of narrow sawfish Anoxypristis cuspidata in Indonesia. Four liters of seawater samples were collected at six locations near the coral reefs ecosystem of Indonesia and filtered at 0.45 μm filter paper. DNA was extracted from the filter paper then Polymerase Chain Reaction (PCR) amplification was performed using the cytochrome c oxidase subunit I (COI) primer and analyzed by Next Generation Sequencing (NGS). The findings revealed that narrow sawfish exist in Indonesian waters, and it also simultaneously showed that environmental DNA could detect rare species. The environmental DNA approach to identifying narrow sawfish can provide reliable results and be used as a survey tool to protect endangered threatened and protected (ETP) species.

scholarly journals Evaluation of Ion Torrent next-generation sequencing for thalassemia diagnosis

2020 ◽  
Vol 48 (12) ◽  
pp. 030006052096777
Author(s):  
Peisong Chen ◽  
Xuegao Yu ◽  
Hao Huang ◽  
Wentao Zeng ◽  
Xiaohong He ◽  
...  
Keyword(s):  
Next Generation Sequencing ◽  
Ion Torrent ◽  
Next Generation ◽  
Large Deletions ◽  
Different Types ◽  
Variant Detection ◽  
Polymerase Chain ◽  
Next Generation Sequencing Ngs ◽  
Accuracy And Repeatability ◽  
Generation Sequencing

Introduction To evaluate a next-generation sequencing (NGS) workflow in the screening and diagnosis of thalassemia. Methods In this prospective study, blood samples were obtained from people undergoing genetic screening for thalassemia at our centre in Guangzhou, China. Genomic DNA was polymerase chain reaction (PCR)-amplified and sequenced using the Ion Torrent system and results compared with traditional genetic analyses. Results Of the 359 subjects, 148 (41%) were confirmed to have thalassemia. Variant detection identified 35 different types including the most common. Identification of the mutational sites by NGS were consistent with those identified by Sanger sequencing and Gap-PCR. The sensitivity and specificities of the Ion Torrent NGS were 100%. In a separate test of 16 samples, results were consistent when repeated ten times. Conclusion Our NGS workflow based on the Ion Torrent sequencer was successful in the detection of large deletions and non-deletional defects in thalassemia with high accuracy and repeatability.

scholarly journals METODE EKSTRAKSI DNA TANAMAN TANPA PRESIPITASI ETANOL UNTUK KEGIATAN POLYMERASE CHAIN REACTION (PCR)

2019 ◽  
Vol 6 (1) ◽  
pp. 29
Author(s):  
Kristianto Nugroho ◽  
Rerenstradika Tizar Terryana ◽  
. Reflinur ◽  
Puji Lestari
Keyword(s):  
Polymerase Chain Reaction ◽  
Dna Extraction ◽  
Simple Sequence Repeat ◽  
Sequence Repeat ◽  
Chain Reaction ◽  
Ethanol Precipitation ◽  
Polymerase Chain ◽  
Next Generation Sequencing Ngs ◽  
Generation Sequencing ◽  
Simple Sequence

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

scholarly journals Ancient human DNA: A history of hype (then and now)

2021 ◽  
pp. 146960532199011
Author(s):  
Elizabeth D Jones ◽  
Elsbeth Bösl
Keyword(s):  
Current Debate ◽  
Chain Reaction ◽  
Hype Cycle ◽  
Human Dna ◽  
Celebrity Status ◽  
History Of ◽  
Polymerase Chain ◽  
And Migration ◽  
Next Generation Sequencing Ngs ◽  
Generation Sequencing

In this article on the history of ancient DNA research, we argue that the innovation of next-generation sequencing (NGS) of the early 2000s has ushered in a second hype cycle much like the first hype cycle the field experienced in the 1990s with the advent of the polymerase chain reaction (PCR). While the first hype cycle centered around the search for the oldest DNA, the field’s current optimism today promotes the rhetoric of revolution surrounding the study of ancient human gnomes. This is evidenced from written sources and personal interviews with researchers who feel the vast amount of data, the conclusions being made from this data, and the ever-increasing celebrity status of the field are perhaps moving too fast for their own good. Here, we use the concept of contamination, in both a literal and figurative understanding of the term, to explore the field’s continuities and disparities. We also argue that a number of additional, figurative interpretations of “contamination” are useful for navigating the current debate between geneticists and archaeologists regarding the origin, evolution, and migration of ancient humans across space and time. Our historical outlook on aDNA’s disciplinary development, we suggest, is necessary to accurately appreciate the state of the field, how it came to be, and where it might go in the future.

scholarly journals A Novel p.G141R Mutation in ILDR1 Leads to Recessive Nonsyndromic Deafness DFNB42 in Two Chinese Han Families

2018 ◽  
Vol 2018 ◽  
pp. 1-6 ◽  
Author(s):  
Xueling Wang ◽  
Longhao Wang ◽  
Hu Peng ◽  
Tao Yang ◽  
Hao Wu
Keyword(s):  
Sanger Sequencing ◽  
Pcr Amplification ◽  
Homozygous Mutation ◽  
Frequent Mutation ◽  
Nonsyndromic Deafness ◽  
Chinese Han ◽  
Targeted Next Generation Sequencing ◽  
Targeted Ngs ◽  
Next Generation Sequencing Ngs ◽  
Generation Sequencing

Genetic hearing impairment is highly heterogeneous. In this study, targeted next-generation sequencing (NGS) in two Chinese Han families identified a novel p.G141R homozygous mutation in ILDR1 as the genetic cause of the deafness. Consistent with the recessive inheritance, cosegregation of the p.G141R variant with the hearing loss was confirmed in members of both families by PCR amplification and Sanger sequencing. SNP genotyping analysis suggested that those two families were not closely related. Our study showed that targeted NGS is an effective tool for diagnosis of genetic deafness and that p.G141R in ILDR1 may be a relatively frequent mutation for DFNB42 in Chinese Hans.

Diagnostik der Sepsis – Teil 2: Erregeridentifikation

2019 ◽  
Vol 54 (01) ◽  
pp. 38-48 ◽  
Author(s):  
Daniel Richter ◽  
Alexandra Heininger ◽  
Karsten Schmidt ◽  
Thomas Schmoch ◽  
Michael Bernhard ◽  
...  
Keyword(s):  
Polymerase Chain Reaction ◽  
Next Generation Sequencing ◽  
Next Generation ◽  
Chain Reaction ◽  
Antimikrobielle Therapie ◽  
Polymerase Chain ◽  
Next Generation Sequencing Ngs ◽  
Generation Sequencing ◽  
Kritisch Kranker

ZusammenfassungIm Rahmen der Sepsis und des septischen Schocks spielen, trotz der zunehmenden Verbreitung von neuen molekularbiologischen Verfahren, der kulturelle Erregernachweis und die Resistenztestung weiterhin die entscheidende Rolle in der antimikrobiellen Therapie auf der Intensivstation. Hierbei kann der Erregernachweis für die antimikrobielle Therapie einerseits direkt aus dem Patientenblut, andererseits aber auch aus diversen anderen Probenmaterialien (respiratorische Sekrete, Punktat, intraoperative Abstriche etc.) geführt werden. Ein Nachteil konventioneller kultureller Verfahren im Kontext kritisch kranker Patienten ist die zeitliche Latenz bis zum Erregernachweis bzw. zum Ergebnis der Resistenztestung. Molekularbiologische Verfahren wie Techniken der Erregerdiagnostik und Resistenztestung, die auf Polymerase Chain Reaction (PCR) oder vor allem Next-Generation Sequencing (NGS) basieren, versprechen hier zwar kürzere Umlaufzeiten, sind aber aktuell noch kein klinischer Standard. Trotzdem besitzen diese Verfahren das Potenzial, einen Paradigmenwechsel in der Erregerdiagnostik herbeizuführen.

scholarly journals Increasing importance of genomics in recognition of infectious animal diseases taking into account OIE requirements

2017 ◽  
Vol 73 (1) ◽  
pp. 10-14
Author(s):  
Woźniakowski Grzegorz. ◽  
Truszczyński Marian ◽  
Pejsak Zygmunt
Keyword(s):  
High Throughput Sequencing ◽  
Animal Health ◽  
Infectious Agents ◽  
Animal Diseases ◽  
Routine Diagnosis ◽  
Animal Pathogens ◽  
Polymerase Chain ◽  
Next Generation Sequencing Ngs ◽  
Generation Sequencing ◽  
World Organisation

Currently, the diagnosis methods applied in bacteriology and virology are frequently focused at modern molecular methods such as polymerase chain reaction (PCR), real-time PCR, loop-mediated isothermal amplification (LAMP) or confirmatory methods for already identified infectious agents of animals such as high throughput sequencing- HTS, or next-generation sequencing (NGS). A number of these methods are officially recommended by the World Organisation for Animal Health (OIE) for the routine diagnosis of major animal diseases with the greatest economical impact. The aim of this paper is to provide to a reader an overview on recent genomic methods used in diagnosis of infectious animal diseases along with their advantages and limitations. These methods facilitate efficient and reliable identification of animal pathogens and allow to characterize and sometimes to identify a newly recognized species of bacteria and viruses.

scholarly journals Environmental DNA (eDNA) metabarcoding: Diversity study around the Pondok Dadap fish landing station, Malang, Indonesia

2019 ◽  
Vol 20 (12) ◽  
Author(s):  
Sapto Andriyono ◽  
MD. JOBAIDUL ALAM ◽  
HYUN-WOO KIM
Keyword(s):  
Marine Fish ◽  
Molecular Identification ◽  
Environmental Dna ◽  
Fish Diversity ◽  
Morphological Identification ◽  
Fish Samples ◽  
Identification Of Species ◽  
Seawater Samples ◽  
Next Generation Sequencing Ngs ◽  
Diversity Study

Abstract. Andriyono S, Jobaidul Alam Md, Kim HW. 2019. Environmental DNA (eDNA) metabarcoding: Diversity study around the Pondok Dadap fish landing station, Malang, Indonesia. Biodiversitas 20: 3772-3781. Molecular identification of species is now fast growing and currently widely applied method in the diversity estimation of aquatic biota; even though morphological identification is still carried out. The molecular approach is beneficial complementing on regular surveys, e.g. use of nets, traps, fishing rods, and even with poisons. In this study, the eDNA metabarcoding was applied to water samples around the Pondok Dadap fish landing station, Indonesia to determine the diversity of fish around the waters and also to identify marine fish landed in this area. Molecular identification was carried out on fish samples obtained from the fish market improved GenBank database on COI and ITS. While, seawater samples were carried out by using the next-generation sequencing (NGS) platform to obtain the eDNA metabarcoding data for the first time. Molecular identification obtained 34 species (68 sequences of COI and ITS regions) belonging to 28 genera, 18 families, 4 orders, while the eDNA metabarcoding approach identified 53 marine fish species by using the MiFish pipeline representing 38 genera, 27 families, and 7 orders. From the present study, we can able to estimated fish diversity by eDNA metabarcoding, and this finding will be helpful for baseline data preparation for future effective management of resources in this area.

More (or less?) bounce for the ounce: a comparison of environmental DNA and classical approaches for bioassessment

2018 ◽  
Vol 69 (6) ◽  
pp. 992 ◽  
Author(s):  
Paul J. McInerney ◽  
Gavin N. Rees
Keyword(s):  
Environmental Dna ◽  
Quantitative Information ◽  
Qualitative Assessment ◽  
Functional Aspects ◽  
The Cost ◽  
Next Generation Sequencing Ngs ◽  
Riparian Plant ◽  
Generation Sequencing ◽  
Selection Of

Next-generation sequencing (NGS) techniques are revolutionising the bioassessment of ecosystems. Herein we use a case study to compare environmental (e)DNA and classical sampling and laboratory identification approaches to assess biotic communities in streams. Both techniques were successful in detecting changes to biotic communities following invasion by a non-native riparian plant. The cost of the eDNA methods was one-sixth that of the classical approach and provided a coarse qualitative assessment of overall eukaryotic structure. Classical macroinvertebrate techniques, although they assess only a subset of eukaryotes, provided high-resolution quantitative information that could be applied to assess functional aspects of the ecosystem. Selection of one method in preference over the other is highly dependent on the nature of the hypothesis to be tested.

scholarly journals TET2Overexpression in Chronic Lymphocytic Leukemia Is Unrelated to the Presence ofTET2Variations

2014 ◽  
Vol 2014 ◽  
pp. 1-6 ◽  
Author(s):  
María Hernández-Sánchez ◽  
Ana Eugenia Rodríguez ◽  
Alexander Kohlmann ◽  
Rocío Benito ◽  
Juan Luis García ◽  
...  
Keyword(s):  
Chronic Lymphocytic Leukemia ◽  
Lymphocytic Leukemia ◽  
Novel Mutations ◽  
Exon Arrays ◽  
Hematopoietic Malignancies ◽  
Healthy Donors ◽  
Polymerase Chain ◽  
Next Generation Sequencing Ngs ◽  
Generation Sequencing

TET2is involved in a variety of hematopoietic malignancies, mainly in myeloid malignancies. Most mutations ofTET2have been identified in myeloid disorders, but some have also recently been described in mature lymphoid neoplasms. In contrast to the large amount of data about mutations ofTET2, some data are available for gene expression. Moreover, the role of TET2 in chronic lymphocytic leukemia (CLL) is unknown. This study analyzes bothTET2expression and mutations in 48 CLL patients.TET2expression was analyzed by exon arrays and quantitative real-time polymerase chain reaction (qRT-PCR). Next-generation sequencing (NGS) technology was applied to investigate the presence ofTET2variations. Overexpression ofTET2was observed in B-cell lymphocytes from CLL patients compared with healthy donors (P= 0.004). In addition, in CLL patients, an overexpression ofTET2was also observed in the clonal B cells compared with the nontumoral cells (P= 0.002). However, no novel mutations were observed. Therefore, overexpression ofTET2in CLL seems to be unrelated to the presence of genomicTET2variations.

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