Search results for: pMYs-GFP Retroviral Vector
#29030821 2017/10/14 Save this To Up
Retroviral Transduction of Quiescent Murine Hematopoietic Stem Cells.Hematopoietic stem cells (HSCs) represent an important target cell population in bone marrow transplantation, cell and gene therapy applications, and the development of leukemia models for research. Because the hematopoietic progeny carries the genetic information of HSCs and replenishes the blood and immune system, corrective gene transfer into HSCs provides an ideal therapeutic approach for many monogenic hematological diseases and a useful tool for studies of HSC function and blood formation in normal and malignant hematopoiesis. However, the efficiency of gene transfer into HSCs has been limited by several features of viral vectors, viral titer, methods of viral transduction, and the property of stem cell quiescence. In this chapter, we describe the production of retrovirus using murine stem cell virus (MSCV)-based retroviral vectors and purification and transduction of murine HSCs.
Macrophage Colony Stimula Macrophage Colony Stimula anti CD34 Hematopoietic p anti CD38 Hematopoietic p Stemez hN2 Human Neuron D 129 Mouse Embryonic Stem Rat Mesenchymal Stem Cell Fontana-Masson Stain Kit Fontana-Masson Stain Kit Anti C Reactive Protein A anti HSV (II) gB IgG1 (mo anti HCMV IE pp65 IgG1 (m
#29017974 2017/10/11 Save this To Up
Vectofusin-1, a potent peptidic enhancer of viral gene transfer forms pH-dependent α-helical nanofibrils, concentrating viral particles.Gene transfer using lentiviral vectors has therapeutic applications spanning from monogenic and infectious diseases to cancer. Such gene therapy has to be improved by enhancing the levels of viral infection of target cells and/or reducing the amount of lentivirus for greater safety and reduced costs. Vectofusin-1, a recently developed cationic amphipathic peptide with a pronounced capacity to enhance such viral transduction, strongly promotes the entry of several retroviral pseudotypes into target cells when added to the culture medium. To clarify the molecular basis of its action the peptide was investigated on a molecular and a supramolecular level by a variety of biophysical approaches. We show that in culture medium vectofusin-1 rapidly forms complexes in the 10 nm range that further assemble into annular and extended nanofibrils. These associate with viral particles allowing them to be easily pelleted for optimal virus-cell interaction. Thioflavin T fluorescence, circular dichroism and infrared spectroscopies indicate that these fibrils have a unique α-helical structure whereas most other viral transduction enhancers form β-amyloid fibrils. A vectofusin-1 derivative (LAH2-A4) is inefficient in biological assays and does not form nanofibrils, suggesting that supramolecular assembly is essential for transduction enhancement. Our observations define vectofusin-1 as a member of a new class of α-helical enhancers of lentiviral infection. Its fibril formation is reversible which bears considerable advantages in handling the peptide in conditions well-adapted to Good Manufacturing Practices and scalable gene therapy protocols.
1297 related Products with: Vectofusin-1, a potent peptidic enhancer of viral gene transfer forms pH-dependent α-helical nanofibrils, concentrating viral particles.Recombinant Viral Antige Recombinant Viral antige Recombinant Viral antige Recombinant Viral antige Recombinant Viral antige Recombinant Viral antige Recombinant Viral antige Recombinant Viral antige Recombinant Viral antige Recombinant Viral antige Recombinant Viral antige Recombinant Viral antige
#28985505 2017/10/06 Save this To Up
Combined CRISPRi/a-Based Chemical Genetic Screens Reveal that Rigosertib Is a Microtubule-Destabilizing Agent.Chemical libraries paired with phenotypic screens can now readily identify compounds with therapeutic potential. A central limitation to exploiting these compounds, however, has been in identifying their relevant cellular targets. Here, we present a two-tiered CRISPR-mediated chemical-genetic strategy for target identification: combined genome-wide knockdown and overexpression screening as well as focused, comparative chemical-genetic profiling. Application of these strategies to rigosertib, a drug in phase 3 clinical trials for high-risk myelodysplastic syndrome whose molecular target had remained controversial, pointed singularly to microtubules as rigosertib's target. We showed that rigosertib indeed directly binds to and destabilizes microtubules using cell biological, in vitro, and structural approaches. Finally, expression of tubulin with a structure-guided mutation in the rigosertib-binding pocket conferred resistance to rigosertib, establishing that rigosertib kills cancer cells by destabilizing microtubules. These results demonstrate the power of our chemical-genetic screening strategies for pinpointing the physiologically relevant targets of chemical agents.
1486 related Products with: Combined CRISPRi/a-Based Chemical Genetic Screens Reveal that Rigosertib Is a Microtubule-Destabilizing Agent.MOUSE ANTI APAAP COMPLEX, B-Phycoerythrin antibody 2-Amino Benzimidazole Su 2-Amino Benzimidazole Su EZH2 KMT6 antibody Isoty Interleukin-34 IL34 (N-t Interleukin-34 IL34 anti CRC3 CD3 (bispecific) Cl 2,3 dinor 6 keto Prostag ROR1 Clone '1B4 antibody RBPMS HERMES Clone '1C12 Glucokinase, islet isofor
#28973449 2017/10/03 Save this To Up
Expression of short hairpin RNAs using the compact architecture of retroviral microRNA genes.Short hairpin RNAs (shRNAs) are effective in generating stable repression of gene expression. RNA polymerase III (RNAP III) type III promoters (U6 or H1) are typically used to drive shRNA expression. While useful for some knockdown applications, the robust expression of U6/H1-driven shRNAs can induce toxicity and generate heterogeneous small RNAs with undesirable off-target effects. Additionally, typical U6/H1 promoters encompass the majority of the ∼270 base pairs (bp) of vector space required for shRNA expression. This can limit the efficacy and/or number of delivery vector options, particularly when delivery of multiple gene/shRNA combinations is required. Here, we develop a compact shRNA (cshRNA) expression system based on retroviral microRNA (miRNA) gene architecture that uses RNAP III type II promoters. We demonstrate that cshRNAs coded from as little as 100 bps of total coding space can precisely generate small interfering RNAs (siRNAs) that are active in the RNA-induced silencing complex (RISC). We provide an algorithm with a user-friendly interface to design cshRNAs for desired target genes. This cshRNA expression system reduces the coding space required for shRNA expression by >2-fold as compared to the typical U6/H1 promoters, which may facilitate therapeutic RNAi applications where delivery vector space is limiting.
1089 related Products with: Expression of short hairpin RNAs using the compact architecture of retroviral microRNA genes.Gryphon™ Retroviral Exp pCAMBIA1301 Vector (gusA pCAMBIA2300 Vector (No Re pCAMBIA2301 Vector (gusA Ofloxacin CAS Number [824 Anti AGO2 Human, Monoclon Anti AGO2 Mouse, Monoclon Expression Media Products Expression Media Products Expression Media Products Anti AGO2 Human, Monoclon Anti AGO2 Mouse, Monoclon
#28962646 2017/09/30 Save this To Up
pIL6-TRAIL-engineered umbilical cord mesenchymal/stromal stem cells are highly cytotoxic for myeloma cells both in vitro and in vivo.Mesenchymal/stromal stem cells (MSCs) are favorably regarded in anti-cancer cytotherapies for their spontaneous chemotaxis toward inflammatory and tumor environments associated with an intrinsic cytotoxicity against tumor cells. Placenta-derived or TRAIL-engineered adipose MSCs have been shown to exert anti-tumor activity in both in-vitro and in-vivo models of multiple myeloma (MM) while TRAIL-transduced umbilical cord (UC)-MSCs appear efficient inducers of apoptosis in a few solid tumors. However, apoptosis is not selective for cancer cells since specific TRAIL receptors are also expressed by a number of normal cells. To overcome this drawback, we propose to transduce UC-MSCs with a bicistronic vector including the TRAIL sequence under the control of IL-6 promoter (pIL6) whose transcriptional activation is promoted by the MM milieu.
1894 related Products with: pIL6-TRAIL-engineered umbilical cord mesenchymal/stromal stem cells are highly cytotoxic for myeloma cells both in vitro and in vivo.Macrophage Colony Stimula Macrophage Colony Stimula MarkerGeneTM in vivo lacZ anti HSV (II) gB IgG1 (mo anti HCMV IE pp65 IgG1 (m anti HCMV gB IgG1 (monocl GLP 1 ELISA Kit, Rat Gluc GLP 2 ELISA Kit, Rat Prog Glucagon ELISA KIT, Rat G Leptin ELISA Kit, Rat Lep CometAssay Electrophoresi Sf9 insect cells
#28950284 2017/09/26 Save this To Up
Overexpression of Insulin-Like Growth Factor-2 in Expanded Endothelial Progenitor Cells Improves Left Ventricular Function in Experimental Myocardial Infarction.Insulin-like growth factors (IGFs) are mediators of growth hormone-induced metabolic and anabolic actions, but also regulate cell growth, differentiation, and apoptosis and show beneficial effects in acute myocardial ischemia. Since endothelial progenitor cells (EPCs) improve myocardial function after acute myocardial infarction, we sought to investigate whether overexpression of IGF-2 in expanded EPCs (eEPCs) further contributes to improvement in left ventricular function after myocardial infarction. EPCs were isolated from human cord blood and transduced by a retroviral vector expression of IGF-2 (IGF-2 eEPCs) or vector only. Expression levels were confirmed by RT-PCR. Vector only-transduced eEPCs or IGF-2 eEPCs were transplanted after ligation of the left anterior descending coronary artery in athymic nude rats. Transplantation of eEPCs improved the left ventricular ejection fraction after 2 weeks. Overexpression of IGF-2 further improved the left ventricular ejection fraction and reduced the myocardial infarction size. Immunohistological analysis revealed an increase in proliferating cells and a decrease in monocytes and apoptotic myocytes within the infarction zone after transplantation of IGF-2-overexpressing eEPCs. Transplantation of IGF-2-overexpressing eEPCs in acute myocardial infarction may improve early myocardial function by enhancing proliferation and limiting the inflammatory response and apoptosis.
2114 related Products with: Overexpression of Insulin-Like Growth Factor-2 in Expanded Endothelial Progenitor Cells Improves Left Ventricular Function in Experimental Myocardial Infarction.Human Insulin-like Growth IGF-1R Signaling Phospho- Insulin promoter factor 1 Human Insulin-like Growth Mouse Insulin-like Growth Rat Insulin-like Growth F IGF1, Insulin-like growth Goat Anti-Human Fibroblas Mouse Anti-Insulin-Like G Human Insulin-like Growth Mouse Insulin-like Growth Rat monoclonal anti mouse
#28931371 2017/09/21 Save this To Up
VISMapper: ultra-fast exhaustive cartography of viral insertion sites for gene therapy.The possibility of integrating viral vectors to become a persistent part of the host genome makes them a crucial element of clinical gene therapy. However, viral integration has associated risks, such as the unintentional activation of oncogenes that can result in cancer. Therefore, the analysis of integration sites of retroviral vectors is a crucial step in developing safer vectors for therapeutic use.
2968 related Products with: VISMapper: ultra-fast exhaustive cartography of viral insertion sites for gene therapy.MOUSE ANTI BOVINE ROTAVIR MarkerGeneTM Fluorescent pCAMBIA1105.1R Vecotr (Gu pCAMBIA1200 Vector (No Re pCAMBIA1300 Vector (No Re pCAMBIA1301 Vector (gusA pCAMBIA2200 Vector (No Re pCAMBIA2301 Vector (gusA MOUSE ANTI CANINE DISTEMP Cellufine Formyl , 50 ml Cellufine Formyl Media Cellufine Formyl , 500 ml
#28930975 2017/09/20 Save this To Up
Streamlined Single Cell TCR Isolation and Generation of Retroviral Vectors for In Vitro and In Vivo Expression of Human TCRs.Although, several methods for sequencing of paired T cell receptor (TCR) alpha and beta chains from single T cells have been developed, none so far have been conducive to downstream in vivo functional analysis of TCR heterodimers. We have developed an improved protocol based on a two-step multiplex-nested PCR, which results in a PCR product that spans entire variable regions of a human TCR alpha and beta chains. By identifying unique restriction sites and incorporating them into the PCR primers, we have made the PCR product compatible with direct sub-cloning into the template retroviral vector. The resulting retroviral construct encodes a chimeric human/mouse TCR with a mouse intracellular domain, which is functional in mouse cells or in in vivo mouse models. Overall, the protocol described here combines human single cell paired TCR alpha and beta chain identification with streamlined generation of retroviral vectors readily adaptable for in vitro and in vivo TCR expression. The video and the accompanying material are designed to give a highly detailed description of the single cell PCR, so that the critical steps can be followed and potential pitfalls avoided. Additionally, we provide a detailed description of the cloning steps necessary to generate the expression vector. Once mastered, the whole procedure from single cell sorting to TCR expression could be performed in a short two-week period.
2535 related Products with: Streamlined Single Cell TCR Isolation and Generation of Retroviral Vectors for In Vitro and In Vivo Expression of Human TCRs.CELLKINES Natural Human I Macrophage Colony Stimula Macrophage Colony Stimula Cultrex In Vitro Angiogen Human integrin aVb3, affi Rabbit Anti-Cell death in Rabbit Anti-Cell death in Human Small Intestine Mic Human Large Intestine Mic Human Internal Mammary Ar GFP Expressing Human Inte Goat Anti-Human Synaptogy
#28917838 2017/09/17 Save this To Up
Human SMOOTHENED inhibits human immunodeficiency virus type 1 infection.Human SMOOTHENED (SMO) was identified by expression cloning as a new host factor that inhibits HIV-1 infection. Forced expression of SMO inhibited HIV-1 replication and infection with a single-round lentiviral vector, but not infection with a murine leukemia virus-based retroviral vector in human MT-4 T cells. Quantitative PCR analyses revealed that stable expression of SMO impaired formation of the integrated form of lentiviral DNA, but did not interrupt reverse transcription. This inhibition was evident in MT-4 and HUT102 human T cell lines expressing low levels of SMO mRNA, but not in SupT1 or Jurkat T cell lines expressing higher levels of SMO mRNA. Depletion of SMO mRNA in Jurkat cells facilitated HIV-1 vector infection, suggesting that endogenous SMO plays a role in limiting lentiviral infection. These results suggest that SMO inhibits HIV-1 replication after completion of reverse transcription but before integration.
1213 related Products with: Human SMOOTHENED inhibits human immunodeficiency virus type 1 infection.Recombinant Human Immunod Recombinant Human Papillo Recombinant Human Papillo HIV 1 p24 Core, Human Imm Type II 5-phosphatase ant Recombinant Human CKMB Ty Recombinant Human CKMB Ty Recombinant Human CKMB Ty Recombinant Human CKMB Ty Recombinant Human CKMM Ty Recombinant Human CKMM Ty Recombinant Human CKMM Ty
#28916331 2017/09/16 Save this To Up
Depressive-like phenotype induced by prenatal dexamethasone in mice is reversed by desipramine.Exposure to prenatal insults has been associated with an increased risk for neuropsychiatric disorders, including depression, but the mechanisms are still poorly understood. Persistent alterations of the HPA axis feedback mechanism as well as adult impaired neurogenesis are believed to play a relevant role in the etiology of depression. In addition, growing evidence points at epigenetic reprogramming as a key factor. We have previously shown that prenatal exposure to the synthetic glucocorticoid dexamethasone (DEX) impairs neurogenesis and leads to late onset of depression-like behavior that does not respond to the SSRI antidepressant fluoxetine (FLX). The aims of this study were to assess the effect of DEX prenatal exposure on the morphology of hippocampal granule neurons and on the expression of genes related to plasticity; and to test whether the SNRI antidepressant desipramine (DMI), unlike FLX, could counteract the effect of prenatal-DEX. C57Bl/6 mice were exposed to DEX (0.05 mg/kg/day) in utero and received intra-hippocampal injection of GFP expressing retroviral vector for labeling of newborn granule cells at eleven months. By twelve months, DEX mice showed depression-like behavior associated with decreased neurogenesis and morphological alterations of the newborn granule cells in the dentate gyrus (DG). Furthermore DEX mice displayed altered expression of genes controlling neurogenesis and neuronal morphology, such as Cdkn1c, p16, TrkB, DISC1 and Reelin. Chronic treatment with DMI led to a significant decrease in immobility time in the forced swim test. In addition, DMI restored neurogenesis, neuronal morphology in the DG, as well as the expression of all related genes. Our results suggest that (1) prenatal DEX induces early and persistent reprogramming effects resulting in altered neurogenesis and neuronal morphology; and (2) DMI treatment reverses DEX-induced depression by restoring the expression of genes relevant to neuronal plasticity.
1577 related Products with: Depressive-like phenotype induced by prenatal dexamethasone in mice is reversed by desipramine.Anti AGO2 Human, Monoclon Anti AGO2 Mouse, Monoclon Anti AGO2 Human, Monoclon Anti AGO2 Mouse, Monoclon BYL-719 Mechanisms: PI3K- Interleukin-34 IL34 (N-t Interleukin-34 IL34 anti HIV1 integrase antibody, Human Epstein-Barr Virus Jurkat Cell Extract (Indu Jurkat Cell Extract (Indu Jurkat Cell Extract (Indu
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