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Biological Procedures Online

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https://www.readbyqxmd.com/read/29881330/method-for-improving-the-quality-of-genomic-dna-obtained-from-minute-quantities-of-tissue-and-blood-samples-using-chelex-100-resin
#1
Utkarsha A Singh, Mukta Kumari, Soumya Iyengar
Background: Although genomic DNA isolation using the Chelex 100 resin is rapid and inexpensive, the DNA obtained by this method has a low concentration in solution and contains suspended impurities. The presence of debris in the DNA solution may result in degradation of DNA on long term storage and inhibition of the polymerase chain reaction. In order to remove impurities and concentrate the DNA in solution, we have introduced modifications in the existing DNA isolation protocol using Chelex-100...
2018: Biological Procedures Online
https://www.readbyqxmd.com/read/29881329/isolation-and-semi-quantitative-pcr-of-na-h-antiporter-sos1-and-nhx-genes-under-salinity-stress-in-kochia-scoparia
#2
Leila Fahmideh, Ziba Fooladvand
Background: Kochia scoparia is a dicotyledonous annual herb and belongs to the Amaranthaceae family. Genetic diversity and resistance to drought stress of this plant has made it widely scattered in different regions which contains highly genetic diversity and great potential as fodder and can grow on salty, drought affected areas. Since the soil salinity has become widely spread, environmental concern has sparked so many debates. An important limiting factor in agricultural production worldwide is the sensitivity of most of the crop to salinity caused by high concentration of salts soil...
2018: Biological Procedures Online
https://www.readbyqxmd.com/read/29743821/blockade-of-itga2-induces-apoptosis-and-inhibits-cell-migration-in-gastric-cancer
#3
Yu-Chang Chuang, Hsin-Yi Wu, Yu-Ling Lin, Shey-Cherng Tzou, Cheng-Hsun Chuang, Ting-Yan Jian, Pin-Rong Chen, Yuan-Ching Chang, Chi-Hsin Lin, Tse-Hung Huang, Chao-Ching Wang, Yi-Lin Chan, Kuang-Wen Liao
Background: Gastric cancer is currently the fourth leading cause of cancer-related death worldwide. Gastric cancer is often diagnosed at advanced stages and the outcome of the treatment is often poor. Therefore, identifying new therapeutic targets for this cancer is urgently needed. Integrin alpha 2 (ITGA2) subunit and the beta 1 subunit form a heterodimer for a transmembrane receptor for extracellular matrix, is an important molecule involved in tumor cell proliferation, survival and migration...
2018: Biological Procedures Online
https://www.readbyqxmd.com/read/29743820/use-of-rna-immunoprecipitation-method-for-determining-sinorhizobium-meliloti-rna-hfq-protein-associations-in-vivo
#4
Mengsheng Gao, Anne Benge, Julia M Mesa, Regina Javier, Feng-Xia Liu
Background: Soil bacterium Sinorhizobium meliloti ( S. meliloti ) forms an endosymbiotic partnership with Medicago truncatula ( M. truncatula ) roots which results in root nodules. The bacteria live within root nodules where they function to fix atmospheric N2 and supply the host plant with reduced nitrogen. The bacterial RNA-binding protein Hfq (Hfq) is an important regulator for the effectiveness of the nitrogen fixation. RNA immunoprecipitation (RIP) method is a powerful method for detecting the association of Hfq protein with specific RNA in cultured bacteria, yet a RIP method for bacteria living in root nodules remains to be described...
2018: Biological Procedures Online
https://www.readbyqxmd.com/read/29618954/treatment-of-advanced-non-small-cell-lung-cancer-with-qi-nourishing-essence-replenishing-chinese-herbal-medicine-combined-with-chemotherapy
#5
Yabin Gong, Zhenye Xu, Changjuan Jin, Haibin Deng, Zhongqi Wang, Weidong Zhou, Ming Zhang, Xiaozhen Zhao, Lifang Wang
Background: To evaluate the effect of qi-nourishing essence-replenishing Chinese herbal medicine combined with chemotherapy in survival of advanced non-small-cell lung cancer(NSCLC) patients with essence and qi deficiency. Methods: A prospective multi-centered randomized controlled study was conducted, and 266 advanced NSCLC patients were enrolled. 126 patients in control group received Vinorelbine plus cisplatin(NP) chemotherapy combined with symptom-oriented Chinese herbs medication(without qi-nourishing essence-replenishing herbs);140 patients in experimental group received NP chemotherapy combined with qi-nourishing essence-replenishing Chinese herbal medication(Kangliu Zengxiao Decoction and modified Feiyanning Decoction, during and after chemotherapy respectively)...
2018: Biological Procedures Online
https://www.readbyqxmd.com/read/29618953/dna-area-and-netosis-analysis-dana-a-high-throughput-method-to-quantify-neutrophil-extracellular-traps-in-fluorescent-microscope-images
#6
Ryan Rebernick, Lauren Fahmy, Christopher Glover, Mandar Bawadekar, Daeun Shim, Caitlyn L Holmes, Nicole Rademacher, Hemanth Potluri, Christie M Bartels, Miriam A Shelef
Background: Neutrophil extracellular traps (NETs), extracellular structures composed of decondensed chromatin and antimicrobial molecules, are released in a process called NETosis. NETs, which are part of normal host defense, have also been implicated in multiple human diseases. Unfortunately, methods for quantifying NETs have limitations which constrain the study of NETs in disease. Establishing optimal methods for NET quantification holds the potential to further elucidate the role of NETs in normal and pathologic processes...
2018: Biological Procedures Online
https://www.readbyqxmd.com/read/29507535/high-sensitivity-and-high-resolution-in-situ-hybridization-of-coding-and-long-non-coding-rnas-in-vertebrate-ovaries-and-testes
#7
Natsumi Takei, Takuma Nakamura, Shohei Kawamura, Yuki Takada, Yui Satoh, Atsushi P Kimura, Tomoya Kotani
Background: Subcellular localization of coding and non-coding RNAs has emerged as major regulatory mechanisms of gene expression in various cell types and many organisms. However, techniques that enable detection of the subcellular distribution of these RNAs with high sensitivity and high resolution remain limited, particularly in vertebrate adult tissues and organs. In this study, we examined the expression and localization of mRNAs encoding Pou5f1/Oct4, Mos, Cyclin B1 and Deleted in Azoospermia-like (Dazl) in zebrafish and mouse ovaries by combining tyramide signal amplification (TSA)-based in situ hybridization with paraffin sections which can preserve cell morphology of tissues and organs at subcellular levels...
2018: Biological Procedures Online
https://www.readbyqxmd.com/read/29507534/silhouette-scores-for-arbitrary-defined-groups-in-gene-expression-data-and-insights-into-differential-expression-results
#8
Shitao Zhao, Jianqiang Sun, Kentaro Shimizu, Koji Kadota
Background: Hierarchical Sample clustering (HSC) is widely performed to examine associations within expression data obtained from microarrays and RNA sequencing (RNA-seq). Researchers have investigated the HSC results with several possible criteria for grouping (e.g., sex, age, and disease types). However, the evaluation of arbitrary defined groups still counts in subjective visual inspection. Results: To objectively evaluate the degree of separation between groups of interest in the HSC dendrogram, we propose to use Silhouette scores...
2018: Biological Procedures Online
https://www.readbyqxmd.com/read/29434528/antitumor-efficacy-of-human-monocyte-derived-dendritic-cells-comparing-effects-of-two-monocyte-isolation-methods
#9
Graça S Marques, Zélia Silva, Paula A Videira
Background: Dendritic cells (DCs), which can be used as anti-cancer vaccines, are generally obtained in vitro from isolated CD14+ monocytes (MoDCs). This generates high cell numbers and allows instructing DCs to guarantee effective antitumor responses. However, the impact of the monocyte isolation step in the antitumor effectiveness of the generated MoDCs is still unknown. Here, we compared the most used immunomagnetic technologies for monocyte isolation: magnetic activated cell sorting (MACS) from Miltenyi Biotec and EasySep from STEM CELL...
2018: Biological Procedures Online
https://www.readbyqxmd.com/read/29434527/visualisation-of-multiple-tight-junctional-complexes-in-human-airway-epithelial-cells
#10
Alysia G Buckley, Kevin Looi, Thomas Iosifidis, Kak-Ming Ling, Erika N Sutanto, Kelly M Martinovich, Elizabeth Kicic-Starcevich, Luke W Garratt, Nicole C Shaw, Francis J Lannigan, Alexander N Larcombe, Graeme Zosky, Darryl A Knight, Paul J Rigby, Anthony Kicic, Stephen M Stick
Background: Apically located tight junctions in airway epithelium perform a fundamental role in controlling macromolecule migration through paracellular spaces. Alterations in their expression may lead to disruptions in barrier integrity, which subsequently facilitates entry of potential bacterial and other pathogens into the host. Furthermore, there is emerging evidence that the barrier integrity of the airway in certain airway inflammatory diseases may be altered. However, there is little consensus on the way this is assessed and measured and the type of cells used to achieve this...
2018: Biological Procedures Online
https://www.readbyqxmd.com/read/29375275/gibson-deletion-a-novel-application-of-isothermal-in-vitro-recombination
#11
Swara Kalva, Jef D Boeke, Paolo Mita
Background: Recombinant DNA technology is today a fundamental tool for virtually all biological research fields. Among the many techniques available for the construction of a "custom DNA" molecule, the isothermal in vitro assembly, or Gibson assembly, allows for an efficient, one-step, scarless recombination-based assembly. Results: Here, we apply and characterize the use of Gibson assembly for the deletion of DNA sequences around a DNA cut. This method, that we named "Gibson Deletion", can be used to easily substitute or delete one or more restriction sites within a DNA molecule...
2018: Biological Procedures Online
https://www.readbyqxmd.com/read/29321719/cancer-chemoprevention-and-therapy-using-chinese-herbal-medicine
#12
REVIEW
Lijing Jiao, Ling Bi, Yan Lu, Qin Wang, Yabin Gong, Jun Shi, Ling Xu
Traditional Chinese medicine (TCM) plays an indispensable role in cancer prevention and treatment. Chinese herbal medicine (CHM) is a key component of TCM and has been practiced for thousands of years. A number of naturally occurring products from Chinese herbs extracts exhibit strong inhibitory properties against carcinogenesis, including CHM single-herb extracts, CHM-derived active components, and CHM formulas (the polyherbal combinations), which regulate JAK/STAT, MAPK, and NF-ҡB pathways. The present review aims to report the cancer-preventive effect of CHM with evidence from cell-line, animal, epidemiological, and clinical experiments...
2018: Biological Procedures Online
https://www.readbyqxmd.com/read/29255379/maintaining-rna-integrity-for-transcriptomic-profiling-of-ex-vivo-cultured-limbal-epithelial-stem-cells-after-fluorescence-activated-cell-sorting-facs
#13
Lei Liu, Frederik Mølgaard Nielsen, Simone Elkjær Riis, Jeppe Emmersen, Trine Fink, Jesper Østergaard Hjortdal, Chris Bath, Vladimir Zachar
Background: Transcriptomic profiling of ex vivo cultured human limbal epithelial stem cells (hLESCs) will foster better understanding of corneal physiology and novel treatment paradigms to limbal stem cell deficiency (LSCD). However, currently such profiling studies are hampered due to difficulties with producing sufficient amounts of intact mRNA for deep RNA sequencing (RNA-seq) from subpopulations sorted on the basis of co-expression of membrane and intracellular antigens by fluorescence-activated cell sorting (FACS)...
2017: Biological Procedures Online
https://www.readbyqxmd.com/read/29151819/production-of-monoclonal-antibodies-against-human-trefoil-factor-3-and-development-of-a-modified-sandwich-elisa-for-detection-of-trefoil-factor-3-homodimer-in-saliva
#14
Saichit Khummuang, Waraporn Phanphrom, Witida Laopajon, Watchara Kasinrerk, Ponlatham Chaiyarit, Supansa Pata
Background: Human trefoil factor (TFF) peptides consist of three members: TFF1, TFF2 and TFF3. TFF3 is the most abundant TFF peptide in saliva. TFF3 homodimer was suggested to be involved in apoptosis inhibition and malignancy. Determination of TFF3 homodimer expression profiles in saliva may lead to new information about oral biology and diseases. The objective of this study was to generate monoclonal antibodies (mAbs) against TFF3 and apply the produced mAbs for the establishment of ELISA for quantification of dimeric TFF3 in saliva...
2017: Biological Procedures Online
https://www.readbyqxmd.com/read/29075153/sv40-large-t-antigen-disrupts-embryogenesis-of-canine-and-porcine-somatic-cell-nuclear-transfer-embryo
#15
Kiyoung Eun, Seon-Ung Hwang, Yeon Woo Jeong, Sunyoung Seo, Seon Yong Lee, Woo Suk Hwang, Sang-Hwan Hyun, Hyunggee Kim
BACKGROUND: Somatic cell nuclear transfer (SCNT) is a useful biotechnological tool for transgenic animal production using genetically modified somatic cells (GMSCs). However, there are several limitations preventing successful transgenic animal generation by SCNT, such as obtaining proper somatic donor cells with a sufficiently long life span and proliferative capacity for generating GMSCs. Here, we established simian virus 40 large T antigen (SV40LT)-mediated lifespan-extended canine fibroblast cells (SV40LT-K9 cells) and evaluated their potential as nuclei donors for SCNT, based on cellular integrity and SCNT embryo development...
2017: Biological Procedures Online
https://www.readbyqxmd.com/read/28959142/enhanced-mutant-screening-in-one-step-pcr-based-multiple-site-directed-plasmid-mutagenesis-by-introduction-of-silent-restriction-sites-for-structural-and-functional-study-of-proteins
#16
Ting-Yu Kuo, Chung-Che Tsai, Hua-Wen Fu
Site-directed mutagenesis (SDM) has been widely used for studying the structure and function of proteins. A one-step polymerase chain reaction (PCR)-based multiple site-directed plasmid mutagenesis method with extended non-overlapping sequence at the 3' end of the primer increases the PCR amplification efficiency and the capacity of multi-site mutagenesis. Here, we introduced silent restriction sites in the primers used in this PCR-based SDM method by utilizing SDM-Assist software to generate mutants of Helicobacter pylori neutrophil-activating protein (HP-NAP), whose gene has low GC content...
2017: Biological Procedures Online
https://www.readbyqxmd.com/read/28932173/efficient-and-inexpensive-transient-expression-of-multispecific-multivalent-antibodies-in-expi293-cells
#17
Xiaotian T Fang, Dag Sehlin, Lars Lannfelt, Stina Syvänen, Greta Hultqvist
BACKGROUND: Immunotherapy is a very fast expanding field within drug discovery and, hence, rapid and inexpensive expression of antibodies would be extremely valuable. Antibodies are, however, difficult to express. Multifunctional antibodies with additional binding domains further complicate the expression. Only few protocols describe the production of tetravalent bispecific antibodies and all with limited expression levels.. METHODS: Here, we describe a protocol that can produce functional tetravalent, bispecific antibodies at around 22 mg protein/l to a low cost...
2017: Biological Procedures Online
https://www.readbyqxmd.com/read/28855851/an-assessment-of-fixed-and-native-chromatin-preparation-methods-to-study-histone-post-translational-modifications-at-a-whole-genome-scale-in-skeletal-muscle-tissue
#18
Sarah-Anne David, Benoît Piégu, Christelle Hennequet-Antier, Maëlle Pannetier, Tiphaine Aguirre-Lavin, Sabine Crochet, Thierry Bordeau, Nathalie Couroussé, Aurélien Brionne, Yves Bigot, Anne Collin, Vincent Coustham
BACKGROUND: Genomic loci associated with histone marks are typically analyzed by immunoprecipitation of the chromatin followed by quantitative-PCR (ChIP-qPCR) or high throughput sequencing (ChIP-seq). Chromatin can be either cross-linked (X-ChIP) or used in the native state (N-ChIP). Cross-linking of DNA and proteins helps stabilizing their interactions before analysis. Despite X-ChIP is the most commonly used method, muscle tissue fixation is known to be relatively inefficient. Moreover, no protocol described a simple and reliable preparation of skeletal muscle chromatin of sufficient quality for subsequent high-throughput sequencing...
2017: Biological Procedures Online
https://www.readbyqxmd.com/read/28814944/cell-counting-and-viability-assessment-of-2d-and-3d-cell-cultures-expected-reliability-of-the-trypan-blue-assay
#19
Filippo Piccinini, Anna Tesei, Chiara Arienti, Alessandro Bevilacqua
BACKGROUND: Whatever the target of an experiment in cell biology, cell counting and viability assessment are always computed. The Trypan Blue (TB) assay was proposed about a century ago and is still the most widely used method to perform cell viability analysis. Furthermore, the combined use of TB with a haemocytometer is also considered the standard approach to estimate cell population density. There are numerous research articles reporting the use of TB assays to compute cell number and viability of 2D and 3D cultures...
2017: Biological Procedures Online
https://www.readbyqxmd.com/read/28785173/an-initial-investigation-of-an-alternative-model-to-study-rat-primordial-germ-cell-epigenetic-reprogramming
#20
Isabelle Hernandez Cantão, Renato Borges Tesser, Taiza Stumpp
BACKGROUND: Primordial germ cells (PGC) are the precursors of the gametes. During pre-natal development, PGC undergo an epigenetic reprogramming when bulk DNA demethylation occurs and is followed by sex-specific de novo methylation. The de novo methylation and the maintenance of the methylation patterns depend on DNA methyltransferases (DNMTs). PGC reprogramming has been widely studied in mice but not in rats. We have previously shown that the rat might be an interesting model to study germ cell development...
2017: Biological Procedures Online
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