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Electroporation

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https://www.readbyqxmd.com/read/29777316/characterization-of-the-complete-sequences-and-stability-of-plasmids-carrying-the-genes-aac-6-ib-cr-or-qnrs-in-shigella-flexneri-in-the-hangzhou-area-of-china
#1
Xiao-Ying Pu, Yaming Gu, Jun Li, Shu-Juan Song, Zhe Lu
The aim of this study was to explore the fluoroquinolone resistance mechanism of aac (6')-Ib-cr and qnrS gene by comparing complete sequences and stability of the aac(6')-Ib-cr- and qnrS-positive plasmids from Shigella isolates in the Hangzhou area of China. The complete sequences of four newly acquired plasmids carrying aac(6')-Ib-cr or qnrS were compared with those of two plasmids obtained previously and two similar reference Escherichia coli plasmids. The results showed that the length, antibiotic resistance genes and genetic environment were different among the plasmids...
May 18, 2018: World Journal of Microbiology & Biotechnology
https://www.readbyqxmd.com/read/29777026/endosomal-tlr3-tlr7-and-tlr8-control-neuronal-morphology-through-different-transcriptional-programs
#2
Yun-Fen Hung, Chiung-Ya Chen, Yi-Chun Shih, Hsin-Yu Liu, Chiao-Ming Huang, Yi-Ping Hsueh
Neuroinflammation is associated with diverse neurological disorders. Endosomal Toll-like receptors (TLRs) including TLR3, TLR7, and TLR8 cell-autonomously regulate neuronal differentiation. However, the mechanisms by which these three TLRs affect neuronal morphology are unclear. In this study, we compare these TLRs in mouse neurons. By combining in vitro neuronal cultures, in utero electroporation, and transcriptomic profiling, we show that TLR8, TLR7, and TLR3 promote dendritic pruning via MYD88 signaling...
May 18, 2018: Journal of Cell Biology
https://www.readbyqxmd.com/read/29776071/effect-of-ac-electric-field-on-the-dynamics-of-a-vesicle-under-shear-flow-in-the-small-deformation-regime
#3
Kumari Priti Sinha, Rochish M Thaokar
Vesicles or biological cells under simultaneous shear and electric field can be encountered in dielectrophoretic devices or designs used for continuous flow electrofusion or electroporation. In this work, the dynamics of a vesicle subjected to simultaneous shear and uniform alternating current (ac) electric field is investigated in the small deformation limit. The coupled equations for vesicle orientation and shape evolution are derived theoretically, and the resulting nonlinear equations are handled numerically to generate relevant phase diagrams that demonstrate the effect of electrical parameters on the different dynamical regimes such as tank treading (TT), vacillating breathing (VB) [called trembling (TR) in this work], and tumbling (TU)...
March 2018: Physical Review. E
https://www.readbyqxmd.com/read/29773754/a-critical-neurodevelopmental-role-for-l-type-voltage-gated-calcium-channels-in-neurite-extension-and-radial-migration
#4
Satoshi Kamijo, Yuichiro Ishii, Shin-Ichiro Horigane, Kanzo Suzuki, Masamichi Ohkura, Junichi Nakai, Hajime Fujii, Sayaka Takemoto-Kimura, Haruhiko Bito
In spite of many association studies linking gene polymorphisms and mutations of L-type Voltage-Gated Ca2+ Channels (VGCC) in neurodevelopmental disorders, such as autism and schizophrenia, specific L-type VGCC roles during brain development remain unclear. Yet, calcium signaling has been shown to be essential for neurodevelopmental processes such as sculpting of neurites, functional wiring and fine tuning of growing networks. To bridge this gap, we performed submembraneous calcium imaging using a membrane-tethered genetically-encoded calcium indicator (GECI) Lck-G-CaMP7...
May 17, 2018: Journal of Neuroscience: the Official Journal of the Society for Neuroscience
https://www.readbyqxmd.com/read/29769736/trna-tracking-for-direct-measurements-of-protein-synthesis-kinetics-in-live-cells
#5
Ivan L Volkov, Martin Lindén, Javier Aguirre Rivera, Ka-Weng Ieong, Mikhail Metelev, Johan Elf, Magnus Johansson
Our ability to directly relate results from test-tube biochemical experiments to the kinetics in living cells is very limited. Here we present experimental and analytical tools to directly study the kinetics of fast biochemical reactions in live cells. Dye-labeled molecules are electroporated into bacterial cells and tracked using super-resolved single-molecule microscopy. Trajectories are analyzed by machine-learning algorithms to directly monitor transitions between bound and free states. In particular, we measure the dwell time of tRNAs on ribosomes, and hence achieve direct measurements of translation rates inside living cells at codon resolution...
May 16, 2018: Nature Chemical Biology
https://www.readbyqxmd.com/read/29769396/bulk-electroporation-mediated-gene-transfer-into-xenopus-tadpole-brain
#6
Cristina Sáenz de Miera, Ethan Parr, Robert J Denver
In vivo gene transfer is a powerful tool for investigating protein function and gene regulation in living organisms. Delivery of plasmid DNA to the brain of Xenopus tadpoles by bulk electroporation-mediated (EM) gene transfer can be used to study the effects of ectopic gene expression on development, physiology, and behavior. It can also be used to mark cells for lineage tracing, investigate the in vivo function of gene regulatory elements when linked to a reporter gene, and introduce mutations into the genome of transfected cells, among other applications...
May 16, 2018: Cold Spring Harbor Protocols
https://www.readbyqxmd.com/read/29761400/transfecting-macrophages
#7
Andrea-Anneliese Keller, Marten B Maeß, Michael Schnoor, Berith Scheiding, Stefan Lorkowski
Transfection is defined as the transfer of foreign nucleic acids into cells. In general, transfection may achieve either overexpression of a gene by the transfer of plasmid DNA or suppression of gene expression by RNA interference after transfer of small interfering RNA. Both approaches allow for the detailed investigation of the function of a particular gene product or mechanisms of gene regulation. Macrophages are considered as hard-to-transfect cells, as they have evolved to recognize foreign nucleic acids and to initiate an immune response to these molecules...
2018: Methods in Molecular Biology
https://www.readbyqxmd.com/read/29760646/dgcr8-promotes-neural-progenitor-expansion-and-represses-neurogenesis-in-the-mouse-embryonic-neocortex
#8
Nadin Hoffmann, Stefan C Weise, Federica Marinaro, Tanja Vogel, Davide De Pietri Tonelli
DGCR8 and DROSHA are the minimal functional core of the Microprocessor complex essential for biogenesis of canonical microRNAs and for the processing of other RNAs. Conditional deletion of Dgcr8 and Drosha in the murine telencephalon indicated that these proteins exert crucial functions in corticogenesis. The identification of mechanisms of DGCR8- or DROSHA-dependent regulation of gene expression in conditional knockout mice are often complicated by massive apoptosis. Here, to investigate DGCR8 functions on amplification/differentiation of neural progenitors cells (NPCs) in corticogenesis, we overexpress Dgcr8 in the mouse telencephalon, by in utero electroporation ( IU Ep)...
2018: Frontiers in Neuroscience
https://www.readbyqxmd.com/read/29760561/an-efficient-system-for-gene-perturbation-in-embryonic-hippocampal-progenitors-using-ex-vivo-electroporation-followed-by-in-vitro-dissociated-cell-culture
#9
Bhavana Muralidharan, Leora D'Souza, Shubha Tole
We established an efficient cell culture assay that permits combinatorial genetic perturbations in hippocampal progenitors to examine cell-autonomous mechanisms of fate specification. The procedure begins with ex vivo electroporation of isolated, intact embryonic brains, in a manner similar to in utero electroporation but with greatly improved access and targeting. The electroporated region is then dissected and transiently maintained in organotypic explant culture, followed by dissociation and plating of cells on coverslips for in vitro culture...
2018: Journal of Experimental Neuroscience
https://www.readbyqxmd.com/read/29760097/the-glycolipid-gm1-reshapes-asymmetric-biomembranes-and-giant-vesicles-by-curvature-generation
#10
Raktim Dasgupta, Markus S Miettinen, Nico Fricke, Reinhard Lipowsky, Rumiana Dimova
The ganglioside GM1 is present in neuronal membranes at elevated concentrations with an asymmetric spatial distribution. It is known to generate curvature and can be expected to strongly influence the neuron morphology. To elucidate these effects, we prepared giant vesicles with GM1 predominantly present in one leaflet of the membrane, mimicking the asymmetric GM1 distribution in neuronal membranes. Based on pulling inward and outward tubes, we developed a technique that allowed the direct measurement of the membrane spontaneous curvature...
May 14, 2018: Proceedings of the National Academy of Sciences of the United States of America
https://www.readbyqxmd.com/read/29758246/creation-of-deletion-and-insertion-clonal-complex-8-staphylococcus-aureus-mutants-using-a-common-cloning-vector
#11
Ping Chen, S L Rajasekhar Karna, Kai P Leung
Deletion and insertion clonal complex 8 Staphylococcus aureus mutants were created without using intermediate host S. aureus RN4220 or temperature-sensitive shuttle vectors. These mutants were created using a common cloning vector by passing the constructs through a modification host and recovering the electroporated cells in a large volume of medium.
May 11, 2018: Journal of Microbiological Methods
https://www.readbyqxmd.com/read/29757586/production-of-recombinant-protein-of-salsola-kali-sal-k1-pollen-allergen-in-lactococcus-lactis
#12
Ziba Ghasemi, Abdol-Reza Varasteh, Malihe Moghadam, Farnaz Sedghi, Farahzad Jabbariazad, Mojtaba Sankian
The Salsola kali pollen is considered the main cause of allergic sensitization in desert and semi-desert regions. We have constructed recombinant Lactococcus lactis producing Sal k1 protein with the aim of using it as a mucosal vaccine for specific immunotherapy. The Sal k1 gene was amplified, and transferred into a PNZ 8148 plasmid. The PNZ8148-Sal k1 recombinant plasmid was transformed into competent E.coli strain MC1061 for replication, and then was isolated and cloned into competent L. lactis by electroporation...
April 2018: Iranian Journal of Allergy, Asthma, and Immunology
https://www.readbyqxmd.com/read/29755766/percutaneous-irreversible-electroporation-with-systemic-treatment-for-locally-advanced-pancreatic-adenocarcinoma
#13
Edward Leen, John Picard, Justin Stebbing, Mark Abel, Tony Dhillon, Harpreet Wasan
Background: The prognosis for unresectable locally advanced pancreatic adenocarcinoma (LAPC) remains poor. There is increasing interest in modern ablative techniques to improve outcomes. We report on the potential value of integrating percutaneous irreversible electroporation (IRE) in patients undergoing systemic chemotherapy. Methods: Seventy-five patients with unresectable pancreatic carcinoma underwent percutaneous IRE after chemotherapy using computerised tomography guidance under general anaesthesia...
April 2018: Journal of Gastrointestinal Oncology
https://www.readbyqxmd.com/read/29753607/maternal-immunization-with-a-dna-vaccine-candidate-elicits-specific-passive-protection-against-post-natal-zika-virus-infection-in-immunocompetent-balb-c-mice
#14
Ran Wang, Xianzheng Liao, Dongying Fan, Lei Wang, Ji Song, Kaihao Feng, Mingyuan Li, Peigang Wang, Hui Chen, Jing An
Zika virus (ZIKV) infection is closely associated in the fetus with microcephaly and in the adults with Guillain-Barré syndrome and even male infertility. It is an urgent international priority to develop a safe and effective vaccine that offers protection to both women of childbearing age and their children. In this study, female immunocompetent BALB/c mice were immunized with a DNA-based vaccine candidate, pVAX1-ZME, expressing the prM/E protein of ZIKV, and the immunogenicity for maternal mice and the post-natal protection for suckling mice were evaluated...
May 9, 2018: Vaccine
https://www.readbyqxmd.com/read/29752690/spatio-temporal-dynamics-of-calcium-electrotransfer-during-cell-membrane-permeabilization
#15
Alexis Guionet, S Moosavi Nejad, Justin Teissié, Takashi Sakugawa, Sunao Katsuki, Hidenori Akiyama, Hamid Hosseini
Pulsed electric fields (PEFs) are applied as physical stimuli for DNA/drug delivery, cancer therapy, gene transformation, and microorganism eradication. Meanwhile, calcium electrotransfer offers an interesting approach to treat cancer, as it induces cell death easier in malignant cells than in normal cells. Here, we study the spatial and temporal cellular responses to 10 μs duration PEFs; by observing real-time, the uptake of extracellular calcium through the cell membrane. The experimental setup consisted of an inverted fluorescence microscope equipped with a color high-speed framing camera and a specifically designed miniaturized pulsed power system...
May 11, 2018: Drug Delivery and Translational Research
https://www.readbyqxmd.com/read/29750170/electroporation-with-cisplatin-against-metastatic-pancreatic-cancer-in-vitro-study-on-human-primary-cell-culture
#16
Olga Michel, Julita Kulbacka, Jolanta Saczko, Justyna Mączyńska, Piotr Błasiak, Joanna Rossowska, Adam Rzechonek
Despite the rapid progression of cancer pharmacotherapy, the high drug resistance of pancreatic ductal adenocarcinoma (PDA) makes it one of the most lethal malignancies. Therefore, there are high expectations associated with experimental therapies, such as electrochemotherapy (ECT). This technique involves the application of short electric pulses to induce transitional permeabilization of the cellular membrane, thus enhancing drug molecules influx. The aim of the study was to investigate the influence of electroporation with cisplatin (CisEP) on the primary culture of human PDA cells from lung metastases-their survival and stress response...
2018: BioMed Research International
https://www.readbyqxmd.com/read/29749639/the-homeodomain-transcription-factor-prox1-is-a-direct-target-of-soxc-proteins-during-developmental-vertebrate-neurogenesis
#17
Anne Jacob, Hannah M Wüst, Johannes M Thalhammer, Franziska Fröb, Melanie Küspert, Simone Reiprich, Elli-Anna Balta, D Chichung Lie, Michael Wegner, Elisabeth Sock
The high-mobility-group-domain containing SoxC transcription factors Sox4 and Sox11 are expressed and required in the vertebrate central nervous system in neuronal precursors and neuroblasts. To identify genes that are widely regulated by SoxC proteins during vertebrate neurogenesis we generated expression profiles from developing mouse brain and chicken neural tube with reduced SoxC expression and found the transcription factor Prox1 strongly downregulated under both conditions. This led us to hypothesize that Prox1 expression depends on SoxC proteins in the developing central nervous system of mouse and chicken...
May 11, 2018: Journal of Neurochemistry
https://www.readbyqxmd.com/read/29748649/efficient-mouse-genome-engineering-by-crispr-ez-technology
#18
Andrew J Modzelewski, Sean Chen, Brandon J Willis, K C Kent Lloyd, Joshua A Wood, Lin He
CRISPR/Cas9 technology has transformed mouse genome editing with unprecedented precision, efficiency, and ease; however, the current practice of microinjecting CRISPR reagents into pronuclear-stage embryos remains rate-limiting. We thus developed CRISPR ribonucleoprotein (RNP) electroporation of zygotes (CRISPR-EZ), an electroporation-based technology that outperforms pronuclear and cytoplasmic microinjection in efficiency, simplicity, cost, and throughput. In C57BL/6J and C57BL/6N mouse strains, CRISPR-EZ achieves 100% delivery of Cas9/single-guide RNA (sgRNA) RNPs, facilitating indel mutations (insertions or deletions), exon deletions, point mutations, and small insertions...
June 2018: Nature Protocols
https://www.readbyqxmd.com/read/29743538/which-technology-to-select-for-primary-focal-treatment-of-prostate-cancer-european-section-of-urotechnology-esut-position-statement
#19
REVIEW
Roman Ganzer, Vinodh Kumar Adithyaa Arthanareeswaran, Hashim U Ahmed, Andrea Cestari, Pascal Rischmann, Georg Salomon, Dogu Teber, Evangelos Liatsikos, Jens-Uwe Stolzenburg, Eric Barret
BACKGROUND: With growing interest in focal therapy (FT) of prostate cancer (PCa) there is an increasing armamentarium of treatment modalities including high-intensity focused ultrasound (HIFU), cryotherapy, focal laser ablation (FLA), irreversible electroporation (IRE), vascular targeted photodynamic therapy (VTP), focal brachytherapy (FBT) and stereotactic ablative radiotherapy (SABR). Currently there are no clear recommendations as to which of these technologies are appropriate for individual patient characteristics...
May 9, 2018: Prostate Cancer and Prostatic Diseases
https://www.readbyqxmd.com/read/29737373/membrane-electroporation-chemical-thermodynamics-and-flux-kinetics-revisited-and-refined
#20
Eberhard Neumann, Sergej Kakorin
The chemical thermodynamic concept for membrane electroporation is critically revisited. The hysteresis in the electric field dependence of the rapid in-field electroporation events (on the in-field hysteresis branch) and the slower post-field pore resealing process (zero-field hysteresis branch) is a typical ensemble property involving rapid single-pore opening-closing events that are temporally and spatially distributed. In the case of spherical membrane shells in homogeneous external fields, the acting local field is dependent on the polar-angular position...
May 8, 2018: European Biophysics Journal: EBJ
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