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Agarose gel

Sandra Vuong, Paul Delgado-Olguin
Genotyping is an invaluable tool for identifying organisms carrying genetic variations including insertions and deletions. This method involves the extraction of DNA from animal tissue samples and subsequent amplification of genomic regions of interest by polymerase chain reaction (PCR). The amplified products are analyzed by agarose gel electrophoresis based on their size.
2018: Methods in Molecular Biology
Zhang Zongze, Yang Xin, Ali Ahmad Awais, Lei Weiqiang, Wang Chunqun, Wenda Di, Zhou Yanqin, Zhao Junlong, Fang Rui, Hu Min
The tetra-primer ARMS-PCR is a rapid, simple and low cost method for single nucleotide polymorphism (SNP) genotyping and has been used to detect SNPs associated with diseases and drug resistance. E198A in the isotype-1 β-tubulin gene is one of the three SNPs associated with benzimidazole resistance in parasitic nematode Haemonchus contortus. However, up to now, only PCR-RFLP method was used to test E198A in H. contortus. In the present study, we developed a tetra-primer ARMS-PCR to detect E198A in H. contortus and the accuracy of the results was compared with that of PCR-coupled sequencing...
March 15, 2018: Veterinary Parasitology
Qiaoxian Huang, Shengke Li, Yuan-Fu Ding, Hang Yin, Lian-Hui Wang, Ruibing Wang
Due to its outstanding capability to facilitate DNA condensation, transportation and endosomal escape, polyethylenimine (PEI) has been frequently studied for gene delivery. However, its molecular weight (M.W.) dependent transfection efficiency and cytotoxicity has severely limited its clinical application. To resolve this dilemma, a supramolecular strategy was developed for the first time, in which PEI with large M.W. (branched, 25 kDa) that has a satisfactory transfection efficiency, yet high non-specific cytotoxicity for gene delivery was wrapped with macrocyclic cucurbit[7]uril (CB[7])...
March 20, 2018: Biomaterials Science
Arun Kumar C Huded, Pavankumar Jingade, Manoj Kumar Mishra
Isolation of high-quality RNA from coffee is challenging because of high level of polysaccharides, polyphenols and other secondary metabolites. In the present study, a rapid and efficient RNA extraction protocol from different tissues of coffee was optimized. Sufficiently high quality and quantity (225.6-454.8 µg/g) of RNA was obtained by using the optimized protocol. The presence of two distinct bands of 28S rRNA and 18S rRNA in agarose gel proved the intactness of the RNA samples. The average spectrophotometric values of the isolated RNA ranged from 1...
March 2018: 3 Biotech
Hao-Hsiang Hsu, John-Kevin Kracht, Laura Elisabeth Harder, Kerstin Rudnik, Gerd Lindner, Katharina Schimek, Uwe Marx, Ralf Pörtner
In vitro cultivated skin models have become increasingly relevant for pharmaceutical and cosmetic applications, and are also used in drug development as well as substance testing. These models are mostly cultivated in membrane-insert systems, their permeability toward different substances being an essential factor. Typically, applied methods for determination of these parameters usually require large sample sizes (e.g., Franz diffusion cell) or laborious equipment (e.g., fluorescence recovery after photobleaching (FRAP))...
February 23, 2018: Journal of Visualized Experiments: JoVE
Radosław Mrówczyński, Artur Jędrzak, Kosma Szutkowski, Bartosz F Grześkowiak, Emerson Coy, Roksana Markiewicz, Teofil Jesionowski, Stefan Jurga
Polydopamine (PDA)-coated magnetic nanoparticles functionalized with mono-6-thio-β-cyclodextrin (SH-βCD) were obtained and characterized by transmission electron microscopy (TEM), Fourier transform infrared spectroscopy (FT-IR), X-ray photoelectron spectroscopy (XPS), Nuclear and Magnetic Resonance Imaging (NMR and MRI), and doxorubicin (DOXO)-loading experiments. The liver cancer cellular internalization of DOXO-loaded nanoparticles was investigated by confocal imaging microscopy. Synthesized nanomaterials bearing a chemotherapeutic drug and a layer of polydopamine capable of absorbing near-infrared light show high performance in the combined chemo- and photothermal therapy (CT-PTT) of liver cancer due to the synergistic effect of both modalities as demonstrated in vitro...
March 16, 2018: Nanomaterials
Carlos German Lemus-Minor, Carmen Cañizares-Nolasco, Maria D D García-Pedrajas, Encarnación Pérez-Artés
Fusarium oxysporum f. sp. dianthi virus 1 (FodV1) was detected in isolate Fod 116 (Fod 116V+ ) of Fusarium oxysporum f. sp. dianthi (Fod), reaching such a high accumulation level that it was clearly visible after agarose gel electrophoresis of total DNA extracts. FodV1 consists of four double-stranded RNA segments, that correspond to a new mycovirus in the Chrysoviridae family. We obtained an isolate of Fod 116 (Fod 116V- ) with only a residual level of FodV1 RNA accumulation by single-conidia selection. Compared to the Fod 116V- , isolate Fod 116V+ showed significant phenotypic alterations in vegetative growth and virulence...
March 8, 2018: Phytopathology
Yuan Huang, Ling Wei, Ai-Min Sun, Bo Li, Cheng-Jun Sun, Wei-Bo Liang, Qiu-Ying Liu, Xiao-Qin Yu, Jing-Yang He, Yang Qin
BACKGROUND: Hepatocellular carcinoma (HCC) is a malignant tumor that severely threatens human health. To date, early detection for HCC patients is particularly significant due to their poor survival rates even after liver resection. METHODS: Therefore, an efficient and sensitive detection method for monitoring liver cancer, multiplex methylation-specific PCR (MSP) coupled with capillary electrophoresis, is developed. RESULTS: Simulations demonstrated that the methylation status of RASSF1A, p16, SFRP1, and ELF could be detected even when DNA equaled or exceeded 12...
March 7, 2018: Journal of Clinical Laboratory Analysis
Allan Rd Nunes, Brenda Elen B Alves, Hannaly Wb Pereira, Yasmin M Nascimento, Ingryd C Morais, José Veríssimo Fernandes, Josélio Mg Araújo, Daniel Cf Lanza
BACKGROUND The genus Flavivirus includes a variety of medically important viruses, including dengue virus (DENV) and Zika virus (ZIKV), which are most prevalent in Brazil. Because the clinical profile of patients affected by different DENV serotypes or ZIKV may be similar, the development of new methods that facilitate a rapid and accurate diagnosis is crucial. OBJECTIVES The current study aimed to develop an improved reverse transcription-polymerase chain reaction (RT-PCR) protocol for universal detection of flaviviruses by using semi-nested primers that discriminate between DENV serotypes and ZIKV...
2018: Memórias do Instituto Oswaldo Cruz
Timothy G Jenkins, Lihua Liu, Kenneth I Aston, Douglas T Carrell
Sperm epigenetic profiles are frequently studied and are of great interest in many fields. One major technical concern when assessing these marks is the potential for somatic cell contamination. Because somatic cells have dramatically different epigenetic signatures, even small levels of contamination can result in significant problems in analysis and interpretation of data. In this study we evaluate an assay, which we designed to offer a reliable 'pre-screen' for somatic cell contamination that directly assesses the DNA being used in the study to determine tissue purity...
April 2018: Systems Biology in Reproductive Medicine
Catherine Lambert, Devang Nijsure, Vincent Huynh, Ryan Wylie
Methods to reversibly control the chemical environment of hydrogels have application in three-dimensional (3D) cell culture to study cell proliferation, migration and differentiation in environments more representative of in vivo environments. Herein, we developed a method to temporally control the chemical environment of agarose hydrogels through non-covalent attachment of peptide motifs. Streptavidin-CGRGDS conjugates were immobilized in desthiobiotin-modified agarose hydrogels through the desthiobiotin-streptavidin interaction (KD 10-11 M)...
March 6, 2018: Biomedical Materials
Katarina Uršič, Tomaž Zupanc, Alja Videtič Paska
Suicide is a well-defined public health problem and is a complex phenomenon influenced by a number of different risk factors, including genetic ones. Numerous studies have examined serotonin system genes. Monoamine oxidase A (MAO-A) is an outer mitochondrial membrane enzyme which is involved in the metabolic pathway of serotonin degradation. Upstream variable number of tandem repeats (uVNTR) in the promoter region of MAOA gene affects the activity of transcription. In the present study we genotyped MAOA-uVNTR polymorphism in 266 suicide victims and 191 control subjects of Slovenian population, which ranks among the European and world populations with the highest suicide rate...
March 2, 2018: Neuroscience Letters
Luděk Vyšín, Tomáš Burian, Egor Ukraintsev, Marie Davídková, Michael E Grisham, Scott Heinbuch, Jorge J Rocca, Libor Juha
In this study, we examined dose-rate effects on strand break formation in plasmid DNA induced by pulsed extreme ultraviolet (XUV) radiation. Dose delivered to the target molecule was controlled by attenuating the incident photon flux using aluminum filters as well as by changing the DNA/buffer-salt ratio in the irradiated sample. Irradiated samples were examined using agarose gel electrophoresis. Yields of single- and double-strand breaks (SSBs and DSBs) were determined as a function of the incident photon fluence...
March 5, 2018: Radiation Research
M J Najafzadeh, V A Vicente, Peiying Feng, A Naseri, Jiufeng Sun, A Rezaei-Matehkolaei, G S de Hoog
The black yeast genus Exophiala includes numerous potential opportunistic species that potentially cause systematic and disseminated infections in immunocompetent individuals. Species causing systemic disease have ability to grow at 37-40 °C, while others consistently lack thermotolerance and are involved in diseases of cold-blooded, waterborne vertebrates and occasionally invertebrates. We explain a fast and sensitive assay for recognition and identification of waterborne Exophiala species without sequencing...
March 5, 2018: Mycopathologia
Maristela P Rangel, Vanessa K de Sá, Tabatha Prieto, João Roberto M Martins, Eloísa R Olivieri, Dirce Carraro, Teresa Takagaki, Vera Luiza Capelozzi
Matrix proteoglycans (PGs) have shown promise as biomarker in malignancies. We employed agarose gel eletrophoresis, quantitative real- time reverse transcription-polymerase chain reaction and immunohistochemistry to evaluate the content of sulfated glicosaminoglycans (chondroitin sulfate and heparan sulfate) and expression of PG (biglycan, glypican, perlecan, syndecan e versican) in patient-matched normal and tumor tissues obtained from resected specimens of lung cancer. A significant increase of heparan sulfate (HS) and chondroitin sulfate (CS) concentrations was found in tumor tissue samples when compared to normal lung tissue samples...
March 3, 2018: Glycoconjugate Journal
Steven Marczak, Katherine Richards, Zeinab Ramshani, Elaine Smith, Satyajyoti Senapati, Reginald Hill, David B Go, Hsueh-Chia Chang
Exosomes carry microRNA biomarkers, occur in higher abundance in cancerous patients than in healthy ones, and because they are present in most biofluids, including blood and urine, can be obtained non-invasively. Standard laboratory techniques to isolate exosomes are expensive, time-consuming, provide poor purity, and recover on the order of 25% of the available exosomes. We present a new microfluidic technique to simultaneously isolate exosomes and preconcentrate them by electrophoresis using a high transverse local electric field generated by ion-depleting ion-selective membrane...
February 27, 2018: Electrophoresis
Ramendra Pal, Subhas C Kundu, Vamsi K Yadavalli
Flexible and thin-film devices are of great interest in epidermal and implantable bioelectronics. The integration of energy storage and delivery devices such as supercapacitors (SCs) with properties of flexibility, miniaturization, biocompatibility, and degradability are sought for such systems. Reducing e-waste, and using sustainable materials and processes are additional desirable qualities. Herein, a silk protein-based biocompatible and degradable thin film micro-supercapacitor (µSC) is reported. A protein carrier with the conducting polymer PEDOT:PSS and reduced graphene oxide dopant, is used as a photopatternable biocomposite ink...
February 26, 2018: ACS Applied Materials & Interfaces
Govindarajan Prasanna, Pu Jing
In this study, chemical chaperone like function of cyanidin-3-O-glucoside (C3G) was investigated through fluorescence spectroscopy, UV-visible spectroscopy, circular dichroism spectroscopy, confocal microscopy, scanning electron microscopy and molecular docking studies. Early and advanced glycation inhibitory effect was evaluated by fluorescence spectroscopy and agarose gel electrophoresis. Amyloids were investigated based on their propensity to bind Congo Red (CR) and Thioflavin T (ThT) by multiple microscopic approaches...
February 20, 2018: Archives of Biochemistry and Biophysics
Ewurama D A Owusu, Samson K Djonor, Charles A Brown, Martin P Grobusch, Petra F Mens
BACKGROUND: Plasmodium falciparum, the most dominant species in sub-Saharan Africa, causes the most severe clinical malaria manifestations. In resource-limited Ghana, where malaria and HIV geographically overlap, histidine-rich protein 2 (HRP2)-based rapid diagnostic test (RDT) is a faster, easier and cheaper alternative to clinical gold standard light microscopy. However, mutations in parasite hrp2 gene may result in missed infections, which have severe implications for malaria control...
February 23, 2018: Malaria Journal
Cansu Ozbayer, Hulyam Kurt, Medine Nur Kebapci, Didem Turgut Cosan, Ertugrul Colak, Hasan Veysi Gunes, Irfan Degirmenci
The vitamin K epoxide reductase complex subunit 1 (VKORC1) gene is expressed in many tissue types, and encodes the VKORC1 protein, which is a key enzyme in the vitamin K cycle. Although researchers have focused on the effects of vitamin K on glucose metabolism, and on its role in the development of type 2 diabetes (T2DM), no consensus has yet been reached. Therefore, here we aimed to investigate the association between VKORC1 variants and the risk of T2DM. The 3673G / A (rs9923231) and 9041G / A (rs7294) VKORC1 variants were investigated by polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) in 100 control individuals and 100 patients with T2DM...
February 22, 2018: International Journal for Vitamin and Nutrition Research. Journal International de Vitaminologie et de Nutrition
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