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FACS replacement

Saixiang Feng, Chenggang Xu, Kaijie Yang, Haihong Wang, Huiying Fan, Ming Liao
In Haemophilus parasuis, the genes HAPS_0217 and HAPS_1695 are predicted to encode long-chain fatty acid-CoA ligases (FACSs). These proteins contain ATP/AMP signature motifs and FACS conserved motifs that are homologous to those in Escherichia coli FadD (EcFadD). In this study, we demonstrate that HAPS_0217 and HAPS_1695 can functionally replace EcFadD in the E. coli fadD mutant JW1794, and were thus designated fadD1 and fadD2, respectively. An evaluation of kinetic parameters indicated that FadD1 and FadD2 have a substrate preference for long-chain fatty acids...
2017: Frontiers in Cellular and Infection Microbiology
Matilda H-C Sheng, Kin-Hing William Lau, Ram Lakhan, Abu Shufian Ishtiaq Ahmed, Charles H Rundle, Patra Biswanath, David J Baylink
The present study was undertaken to determine the mechanism whereby calcitropic hormones and mesenchymal stem cell progeny changes are involved in bone repletion, a regenerative bone process that restores the bone lost to calcium deficiency. To initiate depletion, weanling mice with a mixed C57BL/6 (75%) and CD1 (25%) genetic background were fed a calcium-deficient diet (0.01%) for 14 days. For repletion, the mice were fed a control diet containing 1.2% calcium for 14 days. Depletion decreased plasma calcium and increased plasma parathyroid hormone, 1,25(OH)2D (calcitriol), and C-terminal telopeptide of type I collagen...
April 1, 2017: Endocrinology
John K Nganga, Christian R Samanamu, Joseph M Tanski, Carlos Pacheco, Cesar Saucedo, Victor S Batista, Kyle A Grice, Mehmed Z Ertem, Alfredo M Angeles-Boza
A series of rhenium tricarbonyl complexes coordinated by asymmetric diimine ligands containing a pyridine moiety bound to an oxazoline ring were synthesized, structurally and electrochemically characterized, and screened for CO2 reduction ability. The reported complexes are of the type Re(N-N)(CO)3Cl, with N-N = 2-(pyridin-2-yl)-4,5-dihydrooxazole (1), 5-methyl-2-(pyridin-2-yl)-4,5-dihydrooxazole (2), and 5-phenyl-2-(pyridin-2-yl)-4,5-dihydrooxazole (3). The electrocatalytic reduction of CO2 by these complexes was observed in a variety of solvents and proceeds more quickly in acetonitrile than in dimethylformamide (DMF) and dimethyl sulfoxide (DMSO)...
March 9, 2017: Inorganic Chemistry
Deborah A Sarkes, Margaret M Hurley, Dimitra N Stratis-Cullum
Peptide capture agents have become increasingly useful tools for a variety of sensing applications due to their ease of discovery, stability, and robustness. Despite the ability to rapidly discover candidates through biopanning bacterial display libraries and easily mature them to Protein Catalyzed Capture (PCC) agents with even higher affinity and selectivity, an ongoing challenge and critical selection criteria is that the peptide candidates and final reagent be selective enough to replace antibodies, the gold-standard across immunoassay platforms...
November 9, 2016: Molecules: a Journal of Synthetic Chemistry and Natural Product Chemistry
Thibaut Mirlicourtois, Tche Hong Loquineau, Pierre Yves Chagnon, Thomas Rondepierre, Serge Mirlicourtois, Jean Christophe Bonnardel
OBJECTIVE: The replacement of a mechanical knee with prosthetic class I foot fitting by an electronic knee with prosthetic class II foot was proposed to a 54-years-old schizophrenic patient, active, with right femoral and left tibial traumatic amputation. This fitting evolution was assessed both quantitatively and qualitatively. OBSERVATIONS/METHOD: Compared evaluation of electronic knee with prosthetic class II foot versus mechanical polycentric knee with prosthetic class I foot (fitting unchanged moreover) in: - functional abilities and quality of life using standardized scores (Functional Ambulation Classification, Houghton, Functional Independence Measurement, B...
September 2016: Annals of Physical and Rehabilitation Medicine
Christine A Marsh, Jenny Browne, Jan Taylor, Deborah Davis
BACKGROUND: The conduct of research regardless of the subject or methods employed brings responsibilities and challenges. These are greater when dealing with sensitive topics and vulnerable groups and therefore researchers must navigate a range of complex issues and make choices in relation to practical, ethical and philosophical concerns. While literature dealing with research methodologies and research design may assist to some degree, it cannot provide a clear pathway or template as each research project must respond to a unique set of circumstances...
February 2017: Women and Birth: Journal of the Australian College of Midwives
Tali Benromano, Chaim G Pick, Joav Merick, Ruth Defrin
Objective.: As individuals with intellectual disability (ID) due to cerebral palsy (CP) are at high risk of experiencing pain, measuring their pain is crucial for adequate treatment. While verbal reports are the gold standard in pain measurements, they may not be sufficient in ID. The aim was to detect behavioral/autonomic responses that may indicate the presence and intensity of pain in individuals with CP and ID, using calibrated stimuli, here for the first time. Subjects...
March 1, 2017: Pain Medicine: the Official Journal of the American Academy of Pain Medicine
Christos S Karamitros, Manfred Konrad
Immunogenicity is one of the most common complications occurring during therapy making use of protein drugs of nonhuman origin. A notable example of such a case is bacterial l-asparaginases (L-ASNases) used for the treatment of acute lymphoblastic leukemia (ALL). The replacement of the bacterial enzymes by human ones is thought to set the basis for a major improvement of antileukemic therapy. Recently, we solved the crystal structure of a human enzyme possessing L-ASNase activity, designated hASNase-3. This enzyme is expressed as an inactive precursor protein and post-translationally undergoes intramolecular processing leading to the generation of two subunits which remain noncovalently, yet tightly associated and constitute the catalytically active form of the enzyme...
September 16, 2016: ACS Chemical Biology
Saki Ito, Sorin V Pislaru, Wern Miin Soo, Runqing Huang, Kevin L Greason, Verghese Mathew, Gurpreet S Sandhu, Mackram F Eleid, Rakesh M Suri, Jae K Oh, Vuyisile T Nkomo
BACKGROUND: The impact of pre-procedural RV size and function on outcomes following transcatheter aortic valve replacement (TAVR) is not well established. The aim of this study was to investigate the role of right ventricular size and function on outcomes following TAVR. METHODS AND RESULTS: Between November 2008 and June 2013, 268 consecutive patients (age 80.5±7.9years, aortic valve area 0.79±0.16cm(2)) with symptomatic severe aortic stenosis (AS) undergoing TAVR at Mayo Clinic were included...
October 15, 2016: International Journal of Cardiology
Susan M Byrne, George M Church
CRISPR/Cas9 nuclease systems can create double-stranded DNA breaks at specific sequences to efficiently and precisely disrupt, excise, mutate, insert, or replace genes. However, human embryonic stem or induced pluripotent stem cells (iPSCs) are more difficult to transfect and less resilient to DNA damage than immortalized tumor cell lines. Here, we describe an optimized protocol for genome engineering of human iPSCs using a simple transient transfection of plasmids and/or single-stranded oligonucleotides. With this protocol, we achieve transfection efficiencies greater than 60%, with gene disruption efficiencies from 1-25% and gene insertion/replacement efficiencies from 0...
2015: Current Protocols in Stem Cell Biology
Hong Guo, Stacy Cooper, Alan D Friedman
The murine Cebpa gene contains a +37 kb, evolutionarily conserved 440 bp enhancer that directs high-level expression to myeloid progenitors in transgenic mice. The enhancer is bound and activated by Runx1, Scl, GATA2, C/EBPα, c-Myb, Pu.1, and additional Ets factors in myeloid cells. CRISPR/Cas9-mediated replacement of the wild-type enhancer with a variant mutant in its seven Ets sites leads to 20-fold reduction of Cebpa mRNA in the 32Dcl3 myeloid cell line. To determine the effect of deleting the enhancer in vivo, we now characterize C57BL/6 mice in which loxP sites flank a 688 bp DNA segment containing the enhancer...
2016: PloS One
Raphael Ledermann, Silvan Strebel, Clara Kampik, Hans-Martin Fischer
UNLABELLED: Analysis of bacterial gene function commonly relies on gene disruption or replacement followed by phenotypic characterization of the resulting mutant strains. Deletion or replacement of targeted regions is commonly achieved via two homologous recombination (HR) events between the bacterial genome and a nonreplicating plasmid carrying DNA fragments flanking the region to be deleted. The counterselection of clones that have integrated the entire plasmid in their genome via a single HR event is crucial in this procedure...
May 2016: Applied and Environmental Microbiology
Wenyan Xie, Hongling Wen, Fulu Chu, Shaofeng Yan, Wenli Xie, Bin Lin, Yuzhen Chen, Zhenmei Li, Guijie Ren, Yanyan Song, Li Zhao, Zhiyu Wang
OBJECTIVE: To investigate the effect of the leucine zipper-like motif between HRA and HRB of the human parainfluenza virus 3 fusion protein on fusion activity. METHODS: Site-directed mutagenesis was utilized to substitute the heptadic residues at 257, 264, 271, 278, 285, 292, and 299 in this motif with alanine. Additionally, 3 middle heptadic leucine residues at 271, 278, and 285 were replaced with alanine singly or in combination. A vaccinia virus-T7 RNA polymerase transient expression system was employed to express the wild-type or mutated fusion (F) proteins...
2015: Intervirology
M A Cleary, R Narcisi, K Focke, R van der Linden, P A J Brama, G J V M van Osch
OBJECTIVE: Total bone marrow-derived mesenchymal stem cell (BMSC) populations differ in their potential to undergo chondrogenesis, with individual BMSCs differing in their chondrogenic capacity. The aim of this study was to explore the use of CD105 as a marker to isolate a chondrogenic subpopulation of BMSCs from the total, heterogeneous population. DESIGN: BMSCs were isolated from patients undergoing total hip replacement and following expansion (Passage 1-Passage 5), CD105 expression was investigated by FACS analysis...
May 2016: Osteoarthritis and Cartilage
Xuan Liu, Lunzhi Yuan, Quan Yuan, Yali Zhang, Kun Wu, Tianying Zhang, Yong Wu, Wangheng Hou, Tengyun Wang, Pingguo Liu, James Wai Kuo Shih, Tong Cheng, Ningshao Xia
On the basis of its close phylogenetic relationship with primates, the development of Tupaia belangeri as an infection animal model and drug metabolism model could provide a new option for preclinical studies, especially in hepatitis virus research. As a replacement for primary human hepatocytes (PHHs), primary tupaia hepatocytes (PTHs) have been widely used. Similar to human serum albumin, tupaia serum albumin (TSA) is the most common liver synthesis protein and is an important biomarker for PTHs and liver function...
May 20, 2016: Experimental Animals
Elisabeth Lang, Christine Zelenak, Matthias Eberhard, Rosi Bissinger, Anand Rotte, Mehrdad Ghashghaeinia, Adrian Lupescu, Florian Lang, Syed M Qadri
BACKGROUND/AIMS: The cyclin-dependent kinase 4 (CDK4) participates in the regulation of apoptosis of nucleated cells by altering transcriptional regulation of genes governing cell proliferation and cell death. Similar to apoptosis of nucleated cells, erythrocytes may enter eryptosis, which is characterized by cell shrinkage and cell membrane scrambling with phosphatidylserine (PS) exposure at the cell surface. As mature erythrocytes lack nuclei, acute stimulation of eryptosis cannot result from altered gene expression...
2015: Cellular Physiology and Biochemistry
Federica Battistin, Gabriele Balducci, Nicola Demitri, Elisabetta Iengo, Barbara Milani, Enzo Alessio
We investigated the reactivity of three Ru(ii) precursors -trans,cis,cis-[RuCl2(CO)2(dmso-O)2], cis,fac-[RuCl2(dmso-O)(dmso-S)3], and trans-[RuCl2(dmso-S)4] - towards the diimine linker 2-(2'-pyridyl)pyrimidine-4-carboxylic acid (cppH) or its parent compound 4-methyl-2-(2'-pyridyl)pyrimidine ligand (mpp), in which a methyl group replaces the carboxylic group on the pyrimidine ring. In principle, both cppH and mpp can originate linkage isomers, depending on how the pyrimidine ring binds to ruthenium through the nitrogen atom ortho (N(o)) or para (N(p)) to the group in position 4...
September 21, 2015: Dalton Transactions: An International Journal of Inorganic Chemistry
Yin Bai, Yanmin Chen, Nan Zhang, Xiaochen Guo, Jingzhuang Zhao, Fuxiang Wang, Pengfei Xu, Qingyan Yuan, Jianying Qi, Wenfei Wang, Deshan Li, Guiping Ren
For a long time, researchers have attempted to replace human plasmaderived immunoglobulin against HBV with recombinant HBV antibodies for therapeutic purposes, but failed to develop the products. One of the reasons may be lack of high throughput antibody screening tool. In this study, we screened an antibody library from immunized subjects by a powerful bacterial display technology. The capacity of the ScFv library was 10(9), 117 individual clones against HBV pre- S1 were initially selected and sequenced, the homology of these clones ranged from 59...
2015: Current Pharmaceutical Biotechnology
Kyle A Hollister, Elizabeth S Conner, Mark L Spell, Kristina Deveaux, Léa Maneval, Michael W Beal, Justin R Ragains
Mild conditions are reported for the hydroxylation of aliphatic C-H bonds through radical translocation, oxidation to carbocation, and nucleophilic trapping with H2O. This remote functionalization employs fac-[Ir(ppy)3] together with Tz(o) sulfonate esters and sulfonamides to facilitate the site-selective replacement of relatively inert C-H bonds with the more synthetically useful C-OH group. The hydroxylation of a range of substrates and the methoxylation of two substrates through 1,6- and 1,7-hydrogen-atom transfer are demonstrated...
June 26, 2015: Angewandte Chemie
Ellen Binder, Dipa Natarajan, Julie Cooper, Rania Kronfli, Mara Cananzi, Jean-Marie Delalande, Conor McCann, Alan J Burns, Nikhil Thapar
OBJECTIVES: Enteric neural stem cells provide hope of curative treatment for enteric neuropathies. Current protocols for their harvesting from humans focus on the generation of 'neurospheres' from cultures of dissociated gut tissue. The study aims to better understand the derivation, generation and composition of enteric neurospheres. DESIGN: Gut tissue was obtained from Wnt1-Cre;Rosa26Yfp/Yfp transgenic mice (constitutively labeled neural crest cells) and paediatric patients...
2015: PloS One
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