6,332 results match your criteria protocol amplification

Zika virus RNA detection in blood donors in São Paulo, Brazil.

Hematol Transfus Cell Ther 2021 May 26. Epub 2021 May 26.

Hospital Israelita Albert Einstein (HIAE), São Paulo, SP, Brazil.

Introduction: The Zika Virus (ZIKV) is a single-stranded RNA genome virus, belonging to the family Flaviviridae, genus Flavivirus. Outbreaks around the world have demonstrated that the presence of asymptomatic viremic blood donors provides an increase in the risk of transfusion transmission (TT) and nucleic acid test (NAT) screening has been proposed to ensure the blood safety. This study implemented an "in-house" method to detect ZIKV RNA in blood sample donations. Read More

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Rapid detection of human origin colistin-resistance genes mcr-1, mcr-3, mcr-8, mcr-10 in clinical fecal samples.

Arch Microbiol 2021 Jun 14. Epub 2021 Jun 14.

Key Laboratory of Molecular Epidemiology of Shenzhen, Shenzhen Center for Disease Control and Prevention, Guangdong Province, No. 8 Longyuan Road, Nanshan district, Shenzhen City, 518055, People's Republic of China.

Plasmid-mediated colistin-resistance genes have been reported in human origin clinical samples worldwide which raises its threats to human infections. Notably, mcr-1, mcr-3, mcr-8, and mcr-10 have been reported isolated directly from clinical samples which creates more seriously threaten to human health than other mcr gene types. A multiplex polymerase chain reaction (Multi-PCR) protocol was developed to detect and genotype mobile colistin-resistance genes (mcr-1, mcr-3, mcr-8, mcr-10) in Enterobacteria for clinical laboratory purposes. Read More

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First report of strains related to the phytoplasma associated with Tanzanian Lethal Decline on on the western coast of Madagascar.

Plant Dis 2021 Jun 9. Epub 2021 Jun 9.

MINAE, DPV, Antananarivo, Madagascar;

Madagascar is a high diversity hotspot in the world, and palms are highly represented with nearly 200 endemic species (Rakotoarinivo et al., 2014). Coconut tree (Cocos nucifera) could have been introduced in Madagascar by Austronesians around AD 400 or 700 (Beaujard, 2011). Read More

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Cellular O-Glycome Reporter/Amplification (CORA): Analytical and Preparative Tools to Study Mucin-Type O-Glycans of Living Cells.

Curr Protoc 2021 Jun;1(6):e142

Office of Biotechnology Products, Center for Drug Evaluation and Research, U.S. Food and Drug Administration, Silver Spring, Maryland.

Mucin-type O-glycosylation (O-glycans, O-glycome) is among the most biologically important post-translational modification in glycoproteins but O-glycan structural diversity and expression are poorly understood due to the inadequacy of current analytical methods. We recently developed a new tool termed cellular O-glycome reporter/amplification (CORA), which uses O-glycan precursors, benzyl-α-GalNAc (Bn-α-GalNAc) or azido-Bn-α-GalNAc (N -Bn-α-GalNAc), as surrogates of protein O-glycosylation. Living cells metabolically convert these precursors to all types of O-GalNAc glycans representative of the cells' capabilities. Read More

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Quantitation of Rabies Virus in Various Bovine Brain Structures.

J Vis Exp 2021 May 22(171). Epub 2021 May 22.

Centro Nacional de Investigación Disciplinaria en Microbiología Animal (CENID-MA), INIFAP;

Bovine paralytic rabies (BPR) is a form of viral encephalitis that is of substantial economic importance throughout Latin America, where it poses a major zoonotic risk. Here, our objective was to utilize a laboratory protocol to determine the relative copy number of the rabies virus (RABV) genome in different bovine brain anatomical structures using quantitative real-time reverse transcription polymerase chain reaction (qRT-PCR). qRT-PCR quantifies the specific number of gene copies present in a sample based on fluorescence emitted after amplification that is directly proportional to the amount of target nucleic acid present in the sample. Read More

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Experimental quantum conference key agreement.

Sci Adv 2021 Jun 4;7(23). Epub 2021 Jun 4.

Institute of Photonics and Quantum Sciences, School of Engineering and Physical Sciences, Heriot-Watt University, Edinburgh EH14 4AS, UK.

Quantum networks will provide multinode entanglement enabling secure communication on a global scale. Traditional quantum communication protocols consume pair-wise entanglement, which is suboptimal for distributed tasks involving more than two users. Here, we demonstrate quantum conference key agreement, a cryptography protocol leveraging multipartite entanglement to efficiently create identical keys between users with up to rate advantage in constrained networks. Read More

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Characteristics of pelvic inflammatory disease caused by sexually transmitted disease - An epidemiologic study.

J Gynecol Obstet Hum Reprod 2021 Jun 1;50(9):102176. Epub 2021 Jun 1.

Department of Obstetrics and Gynecology, Meir Medical Center, Kfar Saba, Israel; affiliated with Sackler Faculty of Medicine, Tel Aviv University, Israel.

Introduction: Pelvic inflammatory disease (PID) is an infection of the upper genital organs, diagnosed by clinical findings. The nucleic acid amplification test (NAAT) identify sexually transmitted (STD) pathogens from endocervical swabs, via real time PCR. This study explored the prevalence of STD detected by NAAT for women with PID. Read More

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A New Method for Sequencing the Mitochondrial Genome by Using Long Read Technology.

Methods Mol Biol 2021 ;2277:331-343

Université Paris-Saclay, INRAE, AgroParisTech, Animal Genetic and Integrative Biology, Université Paris-Saclay, Jouy-en-Josas, France.

We describe a protocol to prepare a multiplexed mtDNA library from a blood sample for performing a long read sequencing of the mitochondrial genome. All steps are carefully described to get a high enrichment of mtDNA relative to total DNA extracted from the blood sample. The obtained mutiplexed library allows the production of long sequence mtDNA reads up to 16. Read More

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January 2021

Closed-tube field-deployable loop-mediated isothermal amplification (LAMP) assay based on spore wall protein (SWP) for the visual detection of Enterocytozoon hepatopenaei (EHP).

J Invertebr Pathol 2021 Jul 30;183:107624. Epub 2021 May 30.

ICAR-Central Institute of Brackishwater Aquaculture, #75 Santhome High Road, MRC Nagar, Chennai, India.

Hepatopancreatic microsporidiosis (HPM) is an infectious shrimp disease caused by the microsporidian Enterocytozoon hepatopenaei (EHP). In recent years, the widespread occurrence of EHP poses a significant challenge to the shrimp aquaculture industry. Early, rapid and accurate diagnosis of EHP infection is very much essential for the control of HPM crop-related losses. Read More

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Association of the CAG repeat polymorphism in mitochondrial polymerase gamma (POLG1) with male infertility: a case-control study in an Algerian population.

Afr J Reprod Health 2021 Feb;25(1):67-75

Laboratoire de biologie moléculaire et cellulaire, Université des frères Mentouri Constantine 1, Constantine, Algérie.

Polymorphisms in the have been speculated to be associated with male infertility. The main objective of our study was to CAG repeat polymorphism in POLG1 gene and male mitochondrial DNA polymerase gamma (POLG) assess the possible association of infertility in Algerian population. Genomic DNA from 89 infertile men and 84 controls was extracted using salting-out method. Read More

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February 2021

What does a non-response to induction chemotherapy imply in high-risk medulloblastomas?

J Neurooncol 2021 Jun 2. Epub 2021 Jun 2.

Department of Pediatrics, Institut d'Hématologie et d'Oncologie pédiatrique, 1 Place Professeur Joseph Renaut, 69008, Lyon, France.

Purpose: High-risk medulloblastomas (HR-MB) may not respond to induction chemotherapy, with either post-induction stable (SD) or progressive disease (PD). There is no consensus regarding their optimal management.

Methods: A retrospective, multicentre study investigated patients with non-responder HR-MB treated according to the PNET HR + 5 protocol (NCT00936156) between 01/01/2009 and 31/12/2018. Read More

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Ultrastaging Using Ex Vivo Sentinel Lymph Node Mapping and One-Step Nucleic Acid Amplification (OSNA) in Gastric Cancer: Experiences of a European Center.

Cancers (Basel) 2021 May 29;13(11). Epub 2021 May 29.

Department of Visceral Surgery, University Hospital Augsburg, 86156 Augsburg, Germany.

Background: In this study, the effectiveness of One-step nucleic acid amplification (OSNA) in combination with ex vivo SLN mapping is compared with conventional histology including immunohistochemistry.

Methods: LNs were retrieved from gastrectomy specimens in an unfixed state. After ex vivo SLN mapping using methylene-blue, LNs were sliced to provide samples for histology and OSNA. Read More

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Rapid Detection of (Blackman) (Coleoptera, Curculionidae) with the Loop-Mediated Isothermal Amplification (LAMP) Method.

Plants (Basel) 2021 May 22;10(6). Epub 2021 May 22.

Department of Agricultural, Food, Environmental and Forestry Science and Technology (DAGRI), Plant Pathology and Entomology Section, University of Florence, Piazzale delle Cascine 28, 50144 Florence, Italy.

The walnut twig beetle is a phloem-boring bark beetle responsible, in association with the ascomycete , for the Thousand Cankers Disease (TCD) of walnut trees. The recent finding of TCD in Europe prompted the development of effective diagnostic protocols for the early detection of members of this insect/fungus complex. Here we report the development of a highly efficient, low-cost, and rapid method for detecting the beetle, or even just its biological traces, from environmental samples: the loop-mediated isothermal amplification (LAMP) assay. Read More

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Taguchi method for the optimization of three loop mediated isothermal amplification procedures for Tritrichomonas foetus detection.

Vet Parasitol 2021 Jul 19;295:109462. Epub 2021 May 19.

INCITAP, CONICET, Facultad de Ciencias Exactas y Naturales, Universidad Nacional de La Pampa, Santa Rosa, La Pampa, Argentina. Electronic address:

Taguchi method was used to optimize loop mediated isothermal amplification tests aimed to amplify segments of the elongation factor 1a1 (tf-ef1a1), the 5,8 ribosomal gene (tf-5,8 r) or the beta tubulin 2 (tf-btub2) from the protozoan parasite Tritrichomonas foetus. L9 orthogonal array and quadratic loss functions that penalize deviations from prediction values revealed the effect of amplification reaction components. Analysis of variance (ANOVA) decomposed the contribution of individual factors to a small Ct. Read More

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In Vivo Tracking of Hematopoietic Stem and Progenitor Cell Ontogeny by Cellular Barcoding.

Methods Mol Biol 2021 ;2308:281-300

Institut Curie, Université PSL, Sorbonne Université, CNRS UMR168, Laboratoire Physico Chimie Curie, Paris, France.

Cellular barcoding is a powerful technique that allows for high-throughput mapping of the fate of single cells, notably hematopoietic stem and progenitor cells (HSPCs) after transplantation. Unique artificial DNA fragments, termed barcodes, are stably inserted into HSPCs using lentiviral transduction, making sure that each individual cell receives a single unique barcode. Barcoded HSPCs are transplanted into sublethally irradiated mice where they reconstitute the hematopoietic system through proliferation and differentiation. Read More

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January 2021

Quantitative Determination and Imaging of Gα Signaling in Live Cells via Split-Luciferase Complementation.

Methods Mol Biol 2021 ;2274:69-78

Institute of Pharmacy, University of Regensburg, Regensburg, Germany.

G Protein-coupled receptors (GPCRs) transduce signals elicited by bioactive chemical agents (ligands), such as hormones, neurotransmitters, or cytokines, across the cellular membrane. Upon ligand binding, the receptor undergoes structural rearrangements, which cause the activation of G proteins. This triggers the activation of signaling cascades involving amplification, which takes place after every stage of the cascade. Read More

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Detection of Tumor DNA in Human Plasma with a Functional PLL-Based Surface Layer and Plasmonic Biosensing.

ACS Sens 2021 May 25. Epub 2021 May 25.

Department of Chemical Sciences, University of Catania, Viale Andrea Doria 6, 95122 Catania, Italy.

Standard protocols for the analysis of circulating tumor DNA (ctDNA) include the isolation of DNA from the patient's plasma and its amplification and analysis in buffered solutions. The application of such protocols is hampered by several factors, including the complexity and time-constrained preanalytical procedures, risks for sample contamination, extended analysis time, and assay costs. A recently introduced nanoparticle-enhanced surface plasmon resonance imaging-based assay has been shown to simplify procedures for the direct detection of tumor DNA in the patient's plasma, greatly simplifying the cumbersome preanalytical phase. Read More

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Refinement of 16S rRNA gene analysis for low biomass biospecimens.

Sci Rep 2021 May 24;11(1):10741. Epub 2021 May 24.

Inserm UMR-S1135, Centre d'Immunologie et des Maladies Infectieuses (CIMI-Paris), 91 bd. de l'Hôpital, 75013, Paris, France.

High-throughput phylogenetic 16S rRNA gene analysis has permitted to thoroughly delve into microbial community complexity and to understand host-microbiota interactions in health and disease. The analysis comprises sample collection and storage, genomic DNA extraction, 16S rRNA gene amplification, high-throughput amplicon sequencing and bioinformatic analysis. Low biomass microbiota samples (e. Read More

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Generation, maintenance, and monitoring of gnotobiotic mice.

STAR Protoc 2021 Jun 11;2(2):100536. Epub 2021 May 11.

Department of Critical Care Medicine, Cumming School of Medicine, University of Calgary, Calgary, AB T2N 4A1, Canada.

Gnotobiology has revolutionized the study of microbiota-host interactions. This protocol explains how to generate, maintain, and monitor gnotobiotic mice. Three monitoring methods are presented and compared: bacterial culture, microscopy to visualize the presence (or absence) of bacteria using Gram staining or DNA staining, and 16S rRNA gene amplification and sequencing. Read More

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Designing, optimization, and validation of whole blood direct T-ARMS PCR for precise and rapid genotyping of complex vertebral malformation in cattle.

BMC Biotechnol 2021 May 22;21(1):36. Epub 2021 May 22.

Animal genetics & Breeding Division, ICAR-Central Institute for Research on Cattle, Meerut, UP, India.

Background: DNA testing in the cattle industry undergoes multiple hurdles. Successful genotyping involves the transportation of samples from the field to the laboratory in a chilled environment followed by DNA extraction, and finally, a specific genotyping protocol is followed. Various researches are focused on overcoming these issues. Read More

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Therapeutic efficacy of artemether-lumefantrine for the treatment of uncomplicated Plasmodium falciparum malaria in four malaria endemic states of India.

Malar J 2021 May 21;20(1):229. Epub 2021 May 21.

Global Malaria Programme, World Health Organization, Geneva, Switzerland.

Background: Malaria is a major public health problem in India and accounts for about 88% of malaria burden in South-East Asia. India alone accounted for 2% of total malaria cases globally. Anti-malarial drug resistance is one of the major problems for malaria control and elimination programme. Read More

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Two high-quality de novo genomes from single ethanol-preserved specimens of tiny metazoans (Collembola).

Gigascience 2021 May;10(5)

Max Planck Institute for Plant Breeding Research, Max Planck Genome-centre Cologne, Carl-von-Linné-Weg 10, 50829 Cologne, Germany.

Background: Genome sequencing of all known eukaryotes on Earth promises unprecedented advances in biological sciences and in biodiversity-related applied fields such as environmental management and natural product research. Advances in long-read DNA sequencing make it feasible to generate high-quality genomes for many non-genetic model species. However, long-read sequencing today relies on sizable quantities of high-quality, high molecular weight DNA, which is mostly obtained from fresh tissues. Read More

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ChromInst: A single cell sequencing technique to accomplish pre-implantation comprehensive chromosomal screening overnight.

PLoS One 2021 20;16(5):e0251971. Epub 2021 May 20.

Department of Research and Development, Yikon Genomics (Suzhou) Company Limited, Suzhou, Jiangsu, PR China.

Next Generation Sequencing (NGS) is a powerful tool getting into the field of clinical examination. Its preliminary application in pre-implantation comprehensive chromosomal screening (PCCS) of assisted reproduction (test-tube baby) has shown encouraging outcomes that improves the success rate of in vitro fertilization. However, the conventional NGS library construction is time consuming. Read More

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Purification and Amplification of DNA from Cellulolytic Bacteria: Application for Biogas Production from Crop Residues.

Methods Mol Biol 2021 ;2290:187-201

Department of Biotechnology, School of Agricultural Sciences and Technology, Chinhoyi University of Technology, Chinhoyi, Zimbabwe.

Polymerase chain reaction (PCR) is a popular molecular tool for detection of bacteria. PCR allows millions of copies of a target segment of DNA to be produced. The DNA is extracted from overnight grown cultures of pure bacterial isolates using either the organo-solvent method or a commercial DNA extraction kit. Read More

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January 2021

Conjunctival Commensal Isolation and Identification in Mice.

J Vis Exp 2021 May 1(171). Epub 2021 May 1.

Department of Medicine, Division of Infectious Diseases, Brigham and Women's Hospital, Harvard Medical School;

The ocular surface was once considered immune privileged and abiotic, but recently it appears that there is a small, but persistent commensal presence. Identification and monitoring of bacterial species at the ocular mucosa have been challenging due to their low abundance and limited availability of appropriate methodology for commensal growth and identification. There are two standard approaches: culture based or DNA sequencing methods. Read More

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Prolonged response to treatment based on cell-free DNA analysis and molecular profiling in three patients with metastatic cancer: a case series.

Ther Adv Med Oncol 2021 24;13:17588359211001538. Epub 2021 Mar 24.

Department of Investigational Cancer Therapeutics, Phase I Clinical Trials Program, The University of Texas MD Anderson Cancer Center, Unit 455, 1515 Holcombe Boulevard, Houston, TX 77030, USA.

Background: Patients with advanced and/or metastatic solid tumors have limited treatment options. Mutations that serve as biomarkers of carcinogenesis can be found in cell-free DNA of patients' plasma. Analysis of circulating tumor DNA (ctDNA) was developed as a non-invasive, cost-effective alternative to tumor biopsy when such biopsy is not technically feasible or it is associated with high risk for complications. Read More

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Strategies for Bacteriophage T5 Mutagenesis: Expanding the Toolbox for Phage Genome Engineering.

Front Microbiol 2021 26;12:667332. Epub 2021 Apr 26.

Université Paris-Saclay, CEA, CNRS, Institute for Integrative Biology of the Cell (I2BC), Gif-sur-Yvette, France.

Phage genome editing is crucial to uncover the molecular mechanisms of virus infection and to engineer bacteriophages with enhanced antibacterial properties. Phage genetic engineering relies mostly on homologous recombination (HR) assisted by the targeted elimination of wild-type phages by CRISPR-Cas nucleases. These strategies are often less effective in virulent bacteriophages with large genomes. Read More

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[Vector management of arbovirosis cases notified in the city of Valencia, Spain (2016-2018).]

Rev Esp Salud Publica 2021 May 10;95. Epub 2021 May 10.

Servicio de Sanidad y Consumo. Ayuntamiento de València. Valencia. España.

Objective: Environmental management of imported arboviruses such as dengue (DENV), Zika (ZIKV) or Chikungunya (CHIKV) is a task of great significance for Public Health since the arrival and establishment of the competent vector Aedes albopictus (Asian tiger mosquito) in numerous cities of our country. There are recent examples of autochthonous transmission of some of these arboviruses in Spain probably linked to undetected and / or unmanaged imported cases.

Methods: Vector management interventions were carried out in the city of Valencia (Spain) by the Health Service of the Valencia City Council between 2016 and 2018. Read More

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The Illumina Sequencing Protocol and the NovaSeq 6000 System.

Methods Mol Biol 2021 ;2242:15-42

Department of Biology, University of Firenze, Firenze, Italy.

The NovaSeq 6000 is a sequencing platform from Illumina that enables the sequencing of short reads with an output up to 6 Tb. The NovaSeq 6000 uses the typical Illumina sequencing workflow based on library preparation, cluster generation by in situ amplification, and sequencing by synthesis. Flexibility is one of the major features of the NovaSeq 6000. Read More

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January 2021

Nonradioactive direct telomerase activity detection using biotin-labeled primers.

J Clin Lab Anal 2021 Jun 7;35(6):e23800. Epub 2021 May 7.

Faculty of Hepato-Pancreato-Biliary Surgery, Chinese PLA General Hospital, Beijing, China.

Background: Telomerase is a ribonucleoprotein enzyme responsible for maintenance of telomere length which expressed in more than 85% of cancer cells but undetectable in most normal tissue cells. Therefore, telomerase serves as a diagnostic marker of cancers. Two commonly used telomerase activity detection methods, the telomerase repeated amplification protocol (TRAP) and the direct telomerase assay (DTA), have disadvantages that mainly arise from reliance on PCR amplification or the use of an isotope. Read More

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