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Biological Procedures Online

Yan Lu, Shulin/Sl Li, Shiguo/Sg Zhu, Yabin/Yb Gong, Jun/J Shi, Ling/L Xu
DNA/RNA methylation plays an important role in lung cancer initiation and progression. Liquid biopsy makes use of cells, nucleotides and proteins released from tumor cells into body fluids to help with cancer diagnosis and prognosis. Methylation of circulating tumor DNA (ctDNA) has gained increasing attention as biomarkers for lung cancer. Here we briefly introduce the biological basis and detection method of ctDNA methylation, and review various applications of methylated DNA in body fluids in lung cancer screening, diagnosis, prognosis, monitoring and treatment prediction...
2017: Biological Procedures Online
Yi Su, Weigui Luo, Xiaofei Chen, Huizhen Liu, Yueqing Hu, Wanhuang Lin, Langtao Xiao
BACKGROUND: Indole-3-acetic acid (IAA) extraction and purification are of great importance in auxin research, which is a hot topic in the plant growth and development field. Solid-phase extraction (SPE) is frequently used for IAA extraction and purification. However, no IAA-specific SPE columns are commercially available at the moment. Therefore, the development of IAA-specific recognition materials and IAA extraction and purification methods will help researchers meet the need for more precise analytical methods for research on phytohormones...
2017: Biological Procedures Online
Nicholas B Whitticar, Elisha W Strahler, Parthiban Rajan, Savas Kaya, Craig S Nunemaker
BACKGROUND: Cells are continuously exposed to changes in their environment. Endocrine systems, in particular, communicate by rhythms and feedback loops. In this study, we developed an automated system to produce such conditions for cultured cells in a precisely timed manner. We utilized a programmable pair of syringe pumps for inflow and a peristaltic pump for outflow to create rhythmic pulses at 5-min intervals in solutions that mimic the endogenous patterns of insulin produced by pancreatic islets as a test case...
2016: Biological Procedures Online
Mujahid Iqbal, Yanfei Tao, Shuyu Xie, Yufei Zhu, Dongmei Chen, Xu Wang, Lingli Huang, Dapeng Peng, Adeel Sattar, Muhammad Abu Bakr Shabbir, Hafiz Iftikhar Hussain, Saeed Ahmed, Zonghui Yuan
Aqueous two-phase system (ATPS) is a liquid-liquid fractionation technique and has gained an interest because of great potential for the extraction, separation, purification and enrichment of proteins, membranes, viruses, enzymes, nucleic acids and other biomolecules both in industry and academia. Although, the partition behavior involved in the method is complex and difficult to predict. Current research shows that it has also been successfully used in the detection of veterinary drug residues in food, separation of precious metals, sewage treatment and a variety of other purposes...
2016: Biological Procedures Online
Paolo Mita, Tenzin Lhakhang, Donghui Li, Daniel J Eichinger, David Fenyo, Jef D Boeke
BACKGROUND: The immunoprecipitation (IP) assay is a valuable molecular biology tool applied across a breadth of fields. The standard assay couples IP to immunoblotting (IP/IB), a procedure severely limited as it is not easily scaled for high-throughput analysis. RESULTS: Here we describe and characterize a new methodology for fast and reliable evaluation of an immunoprecipitation reaction. FLIP (FLuorescence IP) relies on the expression of the target protein as a chromophore-tagged protein and couples IP with the measurement of fluorescent signal coating agarose beads...
2016: Biological Procedures Online
Marta Pokrywczynska, Daria Balcerczyk, Arkadiusz Jundzill, Maciej Gagat, Monika Czapiewska, Tomasz Kloskowski, Maciej Nowacki, Agata M Gastecka, Magdalena Bodnar, Alina Grzanka, Andrzej Marszalek, Tomasz Drewa
BACKGROUND: A key requirements for therapy utilizing the tissue engineering methodologies is use of techniques which have the capability to yield a high number of cells, from small tissue biopsy in a relatively short time. Up to date there was no optimal methods of isolation and expansion of urinary bladder smooth muscle cells (UB-SMCs). The aim of this study was to compare isolation and expansion techniques of UB-SMCs to select the most repeatable and efficient one. METHOD: Five protocols of porcine UB- SMCs isolation including enzymatic and explant techniques and three expansion techniques were compared...
2016: Biological Procedures Online
Emily Bliss, Wendy E Heywood, Malika Benatti, Neil J Sebire, Kevin Mills
BACKGROUND: The skin is the largest organ of the human body and is the first line barrier defence against trauma, microbial infiltration and radiation. Skin diseases can be a result of multi-systemic disease or an isolated condition. Due to its proteolysis resistant properties there are relatively few human skin proteomic datasets published compared with other human organs or body fluids. Skin is a challenging tissue to analyse using traditional proteomic techniques due to its high lipid content, insolubility and extensive cross-linking of proteins...
2016: Biological Procedures Online
Ranjita Sengupta, Chandreyee Mukherjee, Nandita Sarkar, Zhihong Sun, Jacob Lesnik, Joseph Huang, Biao Lu
BACKGROUND: A number of integrase defective lentiviral (IDLV) packaging systems have been developed to produce integration deficient lentiviruses for gene delivery and epichromosomal expression. However, despite their growing demand, a comparative study to systemically evaluate the performance efficiency of different mutants on virus packaging and gene expression has not been done. RESULTS: Site-directed mutagenesis was used to generate five integrasedeficient mutants for non-integrative lentiviral packaging (NILVP)...
2016: Biological Procedures Online
Maria Hernandez-Valladares, Elise Aasebø, Olav Mjaavatten, Marc Vaudel, Øystein Bruserud, Frode Berven, Frode Selheim
BACKGROUND: Satisfactory sample preparation for mass spectrometry-based analysis is a critical step in the proteomics workflow. The quality and reproducibility of sample preparation can determine the coverage and confidence of proteomics results. Up to date, several methodologies have been described to produce suitable peptides for mass spectrometry analysis, followed by strategies for enrichment of post-translational modified peptides, if desired. Among them, the filter-aided sample preparation (FASP) has been introduced as a method to allow for removal of denaturants, reductants, alkylators, lipids and nucleic acids prior to trypsin digestion...
2016: Biological Procedures Online
Shulin Li
No abstract text is available yet for this article.
2016: Biological Procedures Online
Russell A Hermansen, Torgeir R Hvidsten, Simen Rød Sandve, David A Liberles
BACKGROUND: The number of species with completed genomes, including those with evidence for recent whole genome duplication events has exploded. The recently sequenced Atlantic salmon genome has been through two rounds of whole genome duplication since the divergence of teleost fish from the lineage that led to amniotes. This quadrupoling of the number of potential genes has led to complex patterns of retention and loss among gene families. RESULTS: Methods have been developed to characterize the interplay of duplicate gene retention processes across both whole genome duplication events and additional smaller scale duplication events...
2016: Biological Procedures Online
Mariana T Pinto, Dimas T Covas, Simone Kashima, Claudia O Rodrigues
BACKGROUND: Endothelial-Mesenchymal-Transition (EndMT) plays an essential role in cardiovascular development, and recently became an attractive therapeutic target based on evidence supporting its involvement in fibrosis and cancer. Important questions that remain to be answered are related to the molecular mechanisms that control EndMT in different organs and distinct pathological conditions. The lack of a detailed protocol for induction of EndMT and the assumption that TGF-β isoforms play similar roles on different types of endothelial cells, limit progress in the field...
2016: Biological Procedures Online
Sushma M Bhosle, Aisha Hunt, Jaideep Chaudhary
BACKGROUND: 2'-5' oligoadenylate synthetases (OAS) are interferon inducible enzymes that polymerizes ATP to 2'-5'-linked oligomers of adenylate (2-5As). As part of the innate immune response, these enzymes are activated by viral double stranded RNA or mRNAs with significant double stranded structure. The 2-5As in turn activate RNaseL that degrade single stranded RNAs. Three distinct forms of OAS exist in human cells (OAS1, 2 and 3) with each form having multiple spliced variants. The OAS enzymes and their spliced variants have different enzyme activities...
2016: Biological Procedures Online
Shaomian Yao
Stem cells are unspecialized/undifferentiated cells that exist in embryos and adult tissues or can be converted from somatic differentiated cells. Use of stem cells for tissue regeneration and tissue engineering has been a cornerstone of the regenerative medicine. Stem cells are also believed to exist in cancer tissues, namely cancer stem cells (CSCs). Growing evidence suggests that CSCs are the culprit of cancer dormancy, progression and recurrence, and thus have recently received great attention. MicroRNAs (miRNAs) are a group of short, non-coding RNAs that regulate expression of a wide range of genes at a post-transcriptional manner...
2016: Biological Procedures Online
Katharina Gimpl, Jessica Klement, Sandro Keller
BACKGROUND: In vitro investigations of membrane proteins usually depend on detergents for protein solubilisation and stabilisation. The amount of detergent bound to a membrane protein is relevant to successful experiment design and data analysis but is often unknown. Triple-detection size-exclusion chromatography enables simultaneous separation of protein/detergent complexes and protein-free detergent micelles and determination of their molar masses in a straightforward and absolute manner...
2016: Biological Procedures Online
Michela Casanova, Lorenzo Pasotti, Susanna Zucca, Nicolò Politi, Ilaria Massaiu, Cinzia Calvio, Maria Gabriella Cusella De Angelis, Paolo Magni
BACKGROUND: Circular plasmid-mediated homologous recombination is commonly used for marker-less allelic replacement, exploiting the endogenous recombination machinery of the host. Common limitations of existing methods include high false positive rates due to mutations in counter-selection genes, and limited applicability to specific strains or growth media. Finally, solutions compatible with physical standards, such as the BioBrick™, are not currently available, although they proved to be successful in the design of other replicative or integrative plasmids...
2016: Biological Procedures Online
Marisa Egan, Jasmine Ramirez, Christian Xander, Chirag Upreti, Shantanu Bhatt
BACKGROUND: The ability to introduce site-specific mutations in bacterial pathogens is essential towards understanding their molecular mechanisms of pathogenicity. This has been greatly facilitated by the genetic engineering technique of recombineering. In recombineering, linear double- or single-stranded DNA molecules with two terminal homology arms are electroporated into hyperrecombinogenic bacteria that express a phage-encoded recombinase. The recombinase catalyzes the replacement of the endogenous allele with the exogenous allele to generate selectable or screenable recombinants...
2016: Biological Procedures Online
Marco Man Kin Tsui, Anri Itoh, Mohamed Amgad, Shao-Fang Wang, Toshio Sasaki
BACKGROUND: One of the most notable recent advances in electron microscopy (EM) was the development of genetically-encoded EM tags, including the fluorescent flavoprotein Mini-SOG (Mini-Singlet Oxygen Generator). Mini-SOG generates good EM contrast, thus providing a viable alternative to technically-demanding methods such as immuno-electron microcopy (immuno-EM). Based on the Mini-SOG technology, in this paper, we describe the construction, validation and optimization of a series of vectors which allow expression of Mini-SOG in the Drosophila melanogaster genetic model system...
2016: Biological Procedures Online
Ashok Kumar, Kartik Dhar, Shamsher Singh Kanwar, Pankaj Kumar Arora
Lipases are industrial biocatalysts, which are involved in several novel reactions, occurring in aqueous medium as well as non-aqueous medium. Furthermore, they are well-known for their remarkable ability to carry out a wide variety of chemo-, regio- and enantio-selective transformations. Lipases have been gained attention worldwide by organic chemists due to their general ease of handling, broad substrate tolerance, high stability towards temperatures and solvents and convenient commercial availability. Most of the synthetic reactions on industrial scale are carried out in organic solvents because of the easy solubility of non-polar compounds...
2016: Biological Procedures Online
Meral Miraloglu, Ergul Belge Kurutas, Perihan Ozturk, Ozer Arican
BACKGROUND: Tinea pedis (TP) is an infection of the feet caused by fungi. The infectious diseases caused by dermatophytes are mainly related to the enzymes produced by these fungi. Up to the now, the local 8-iso-prostaglandin F2α (8-iso-PGF2α), concentration as oxidative stress biomarker and trace elements status have not been published in patients with TP. The aim of this study is to evaluate the relationship between oxidative stress and trace elements (Cu, Zn, Se), and to evaluate the ratios of Cu/Zn and Cu/Se in this disorder...
2016: Biological Procedures Online
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