At least 10% of spontaneous chromosomal antibiotic resistant mutants of bacteria express a strain-dependent graded reduction of virulence; this correlates linearly with a prolonged generation time. Occasionally, these mutants are temperature sensitive or/and auxotrophe. The work described in this paper provides evidence that in such strains the resistance and the accompanying markers exist only as a functional genetic unit. In a series of transduction experiments with a pathogenic strain of Salmonella typhimurium, it was found that without exception, the resistance and the additional markers were 100% simultaneously transferred. Furthermore, antibiotic-resistant Escherichia coli mutants with prolonged generation time, were isolated from faecal samples; it is thus indicated that, such innocuous mutants occur at any time in the intestine. It is concluded that concerns connecting such mutants to the possibility of resistance dissemination are unfounded; furthermore, even if transfer of resistance ...
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The transduction efficiency can be measured by assessing the fraction of cells expressing TurboGFP or TurboRFP after transduction. This can be achieved by simple microscopic observation using a fluorescence microscope or by FACS analysis 48-72 hours after transduction ...
Finden Sie alle Bücher von Michael D. Mendenhall, Robert C. Dickson - Signal Transduction Protocols. Bei der Büchersuchmaschine eurobuch.com können Sie antiquarische und Neubücher VERGLEICHEN UND SOFORT zum Bestpreis bestellen. 1617374482
Specialized transduction occurs only in some temperate phages. But specialized transduction is an extremely efficient gene transfer mechanism.. In some occasions, DNA from a specific region of the host chromosome is integrated directly into the virus genome-usually replacing some viral genes. The resulting defective transducing phage (temperate phage) particles now have bacterial DNA as a part of genome.. To understand the process of specialized transduction, you must first be aware about lytic cycle of Bacteriophage.. Lets contrast between normal lysogenic cycle and mechanism of transduction. ...
Large collection of high quality biology pictures, photos, images, illustrations, diagrams and posters on marine biology, cell biology, microbiology... for educational purposes.. ...
Large collection of high quality biology pictures, photos, images, illustrations, diagrams and posters on marine biology, cell biology, microbiology... for educational purposes.. ...
The Determination of Importance of Sequences Neighboring the Psi Sequence in Lentiviral Vector Transduction and Packaging Efficiency. . Biblioteca virtual para leer y descargar libros, documentos, trabajos y tesis universitarias en PDF. Material universiario, documentación y tareas realizadas por universitarios en nuestra biblioteca. Para descargar gratis y para leer online.
TY - JOUR. T1 - Bioengineered AAV Capsids with Combined High Human Liver Transduction In Vivo and Unique Humoral Seroreactivity. AU - Paulk, Nicole K.. AU - Pekrun, Katja. AU - Zhu, Erhua. AU - Nygaard, Sean. AU - Li, Bin. AU - Xu, Jianpeng. AU - Chu, Kirk. AU - Leborgne, Christian. AU - Dane, Allison P.. AU - Haft, Annelise. AU - Zhang, Yue. AU - Zhang, Feijie. AU - Morton, Chris. AU - Valentine, Marcus B.. AU - Davidoff, Andrew M.. AU - Nathwani, Amit C.. AU - Mingozzi, Federico. AU - Grompe, Markus. AU - Alexander, Ian E.. AU - Lisowski, Leszek. AU - Kay, Mark A.. PY - 2017. Y1 - 2017. N2 - Existing recombinant adeno-associated virus (rAAV) serotypes for delivering in vivo gene therapy treatments for human liver diseases have not yielded combined high-level human hepatocyte transduction and favorable humoral neutralization properties in diverse patient groups. Yet, these combined properties are important for therapeutic efficacy. To bioengineer capsids that exhibit both unique seroreactivity ...
We and others have now shown that transduced primary Ag-specific CD4+ T cells provide many beneficial features for targeted immunotherapy (58). Retrovirally transduced cells were easily selected for dose of transgene based on reporter gene expression and were then safely transferred to recipient animals for local product delivery. Although pGC retroviral vectors use a constitutive promoter, in vitro analysis has demonstrated that transgene expression correlates with the activation state of the CD4+ T cell (G. L. Costa and J. M. Benson, unpublished observations). Therefore, it is reasonable to conclude that transduced T cells require Ag presentation in vivo to produce transgenic proteins. This was evident in bioluminescent cell trafficking studies (Fig. 4⇑A), in which luciferase production was no longer detected in naive mice once transduced T cells returned to a quiescent state (9 days after transfer). Interestingly, we observed that MBP-specific hybridomas were more efficient in reducing EAE ...
A mathematical model for nonrandom generalized transduction is proposed and analyzed. The model takes into account the finite number of transducing particle classes for any given marker. The equations for estimation of the distance between markers from cotransduction frequency data are derived and standard errors of the estimates are given. The obtained relationships depend significantly on the number of classes of transducing fragments. The model was applied to estimate the number of transducing fragment classes for a given marker in transduction with phage P22 of Salmonella typhimurium. It was found that the literature data on frequencies of cotransduction in crosses with mutual substitution of selective and nonselective markers can be rationalized most accurately by assuming that the mean number of classes is equal to 2. An improved method for analysis of cotransduction data is proposed on the basis of our model and the results of calculation. The method relies on solving a set of algebraic ...
Investigation of the role of target cell factors in retrovirus transduction. . Biblioteca virtual para leer y descargar libros, documentos, trabajos y tesis universitarias en PDF. Material universiario, documentación y tareas realizadas por universitarios en nuestra biblioteca. Para descargar gratis y para leer online.
Because of the increased efficiency of transduction and expression, and the possible decreased potential for adverse effects, such as insertional mutagenesis compared to retroviruses, we used lentiviral-vector technology to express the fusion constructs in primary human T cells using clinically validated techniques (15). The cDNA sequences containing the various fusion constructions were cloned into a third-generation lentiviral vector in which the CMV promoter was replaced with the EF-1α promoter (16). Lentiviral vector supernatants transduced primary T cells with high efficiency (Fig. 1B). In preliminary experiments, the SS1-transduced T cells were found to have sustained proliferation in the presence of various cell lines that express mesothelin, while culture in the absence of mesothelin-expressing feeder cells failed to sustain T-cell proliferation (data not shown). All constructs were brightly expressed on the surface of CD4 and CD8 T cells, and expression was stable for at least 2 months ...
Generalized transduction is commonly used to move transposon-induced mutations among bacterial strains by selecting for inheritance of a transposonencoded resistance determinant. Although complete cotransduction of the resistance determinant and the chromosomal mutation might be expected, it is often found that when Tn5(Kan) insertion mutations are transduced by bacteriophage P1 most of the nonmutant kanamycin-resistant transductants are due to specialized transduction of Tn5. Such P1::Tn5 specialized transducing phage are not found when a mutant Tn5 element lacking a functional transposase is employed.. ...
... Determine retroviral transduction efficiency and gene expres...We describe three new ViraPort retroviral reporter vectors: pFB-hrGFP ...Dispensing with marker g...,Versatile,Reporter,Vectors,for,Monitoring,Viral,Transduction,biological,advanced biology technology,biology laboratory technology,biology device technology,latest biology technology
Retroviral (MLV-based) and lentiviral (HIV-1-based) vectors were harvested once at 1.5 days and again at 2.5 days post-transfection. Retroviral genomic particles pseudotyped with VSVGs (gMLV-VSVG) were produced at a higher titer during the first harvest than during the second harvest (Fig. 1A). However, more retroviral transducing particles (functional ones, tMLV-VSVG) were produced during the second harvest than during the first harvest. The number of gMLV-VSVG particles required for the transduction of a single cell (genomic to transducing particle ratio) can be calculated to be 2,200 for the first harvest, but 1,200 for the second harvest. This result indicates that gMLV-VSVG particles assembled at later time points post-transfection are more infectious than the ones assembled earlier.. In contrast, both lentiviral genomic and transducing particles pseudotyped with VSVGs (gHIV1-VSVG and tHIV1-VSVG, respectively) were produced at a higher titer during the first harvest than during the second ...
Background. Gene therapy with LentiGlobin Drug Product (DP), which contains autologous CD34+ hematopoietic stem cells transduced with the betibeglogene darolentivec (BB305) lentiviral vector, has been shown to eliminate red blood cell (RBC) transfusions in most patients with TDT with non-β0/β0 genotypes in the prior HGB-204 (Northstar) phase 1/2 study, with a safety profile consistent with myeloablative conditioning. A key finding in HGB-204 was that the average number of therapeutic gene copies per CD34+ cell (i.e. vector copy number [VCN] per diploid genome; median 0.7, range 0.3-1.5) in the DP correlated with peripheral HbAT87Q (transgenic hemoglobin [Hb]) expression at 6 months. To optimize the proportion of patients able to achieve "transfusion independence" and increase unsupported Hb levels after treatment, a refined manufacturing process for LentiGlobin DP was used, to increase the DP VCN and the proportion of transduced cells. Herein, we report initial data from the phase 3 HGB-207 ...
Inhibitory effect of adenovirus-uteroglobin transduction on the growth of lung cancer cell lines. - Jae Cheol Lee, Kyung-Ho Park, Seon Jin Han, Chul-Gyu Yoo, Choon-Taek Lee, Sung Koo Han, Young-Soo Shim, Young Whan Kim
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Our Human Fibroblast Immortalization Kits include an Immortalization Reagent and Virus Transduction Enhancer, increasing transduction rate up to 10-fold.
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Do you know if lentivirus based vectors (allowing stable transduction of cells) insert to host DNA at specific sequences or randomly? I know it is a replication independent process ...
The Signal Transduction and Therapeutics Program Area is composed of 37 members, spanning 16 Departments within UCLA. In the past competing cycle, investigators...
The Cen-ters for Dis-ease Con-trol and Pre-ven-tion plans to scale back or dis-con-tinue work to pre-vent in-fec-tious-dis-ease epi-demics, other health threats in 39 for-eign coun-tries be-cause it ex-pects fund-ing to end, the agency told em-ploy-ees. https://t.co/ ...
The present study was undertaken to analyse the capability of HIV-1 derived TAT protein transduction domain (PTD) fused with Green Fluorescent Protein (TAT-GFP) as a delivery vehicle into a range of protozoan parasites. Successful transduction of native purified TAT-GFP was observed by fluorescent microscopy in Cryptosporidium parvum, Giardia duodenalis, and Neospora caninum. The ability to transduce peptides and other cargo into protozoan parasites, will greatly assist in the delivery of future peptide-based drugs and target validation peptides.. ...
The effect of chemotherapy drug Mitomycin C (MMC) in combination with recombinant adeno-associated virus II (rAAV2) in cancer therapy was investigated, and the mechanism of MMC affecting rAAV2s bioactivity was also studied. The combination effect was evaluated by the level of GFP and TNF expression in a human glioma cell line, and the mechanism of MMC effects on rAAV mediated gene expression was investigated by AAV transduction related signal molecules. C57 and BALB/c nude mice were injected with rAAV-EGFP or rAAV-TNF alone, or mixed with MMC, to evaluate the effect of MMC on AAV-mediated gene expression and tumor suppression. MMC was shown to improve the infection activity of rAAV2 both in vitro and in vivo. Enhancement was found to be independent of initial rAAV2 receptor binding stage or subsequent second-strand synthesis of target DNA, but was related to cell cycle retardation followed by blocked genome degradation. In vivo injection of MMC combined with rAAV2 into the tumors of the animals
Lenti-X Accelerator is a magnetic bead-based technology designed to accelerate lentiviral transduction and MMLV retrovirus transduction experiments. Lenti-X Accelerator allows you to complete this process in only 30 minutes, without Polybrene.
Lenti-X Accelerator is a magnetic bead-based technology designed to accelerate lentiviral transduction and MMLV retrovirus transduction experiments. Lenti-X Accelerator allows you to complete this process in only 30 minutes, without Polybrene.
A recombinant adeno-associated virus serotype 2 (rAAV2) vector has been altered to carry the human MERTK (hMERTK) gene. This vector has been shown to restore vision in animal models that resemble human MERTK-associated Retinitis Pigmentosa (RP), an incurable retinal degeneration that causes severe vision loss. The proposed study is an open label, Phase I clinical trial of subretinal rAAV2-VMD2-hMERTK administration to individuals with MERTK-associated retinal disease. This trial will lead to a greater understanding of the safety and thereby potential value of gene transfer in MERTK-associated retinal disease and will have implications for other forms of retinal degenerative disease amenable to this type of intervention ...
A recombinant adeno-associated virus serotype 2 (rAAV2) vector has been altered to carry the human MERTK (hMERTK) gene. This vector has been shown to restore vision in animal models that resemble human MERTK-associated Retinitis Pigmentosa (RP), an incurable retinal degeneration that causes severe vision loss. The proposed study is an open label, Phase I clinical trial of subretinal rAAV2-VMD2-hMERTK administration to individuals with MERTK-associated retinal disease. This trial will lead to a greater understanding of the safety and thereby potential value of gene transfer in MERTK-associated retinal disease and will have implications for other forms of retinal degenerative disease amenable to this type of intervention ...
Recombinant Adeno-associated virus 2 ATCC ® VR-1616™ Designation: Recombinant Adeno-associated Virus 2 Reference Standard Stock (rAAV2 RSS) Application:
Recombinant Adeno-associated virus 2 ATCC ® VR-1616™ Designation: Recombinant Adeno-associated Virus 2 Reference Standard Stock (rAAV2 RSS) Application:
TY - JOUR. T1 - G2 Cell Cycle Arrest and Cyclophilin A in Lentiviral Gene Transfer. AU - Zhang, Shangming. AU - Joseph, Guiandre. AU - Pollok, Karen. AU - Berthoux, Lionel. AU - Sastry, Lakshmi. AU - Luban, Jeremy. AU - Cornetta, Kenneth. PY - 2006/10. Y1 - 2006/10. N2 - Lentiviral vectors derived from the human immunodeficiency virus-1 (HIV-1) have a higher propensity to transduce nondividing cells compared to vectors based on oncoretroviruses. We report here that genistein, a previously known protein tyrosine kinase (PTK) inhibitor and G2 cell cycle arrest inducer, significantly enhanced lentiviral transduction in a dose-dependent manner. Increased transduction, as measured by vector expression, was seen in a variety of human cell lines, murine primary lymphocytes, and primary human CD34+ peripheral blood progenitor cells as well. Increased vector expression was also associated with an increase in vector DNA copy number, as assessed by quantitative PCR. Genistein-mediated G2 cell cycle arrest, ...
Because the features and kinetics of adeno−associated virus(AAV)−mediated gene transfer to endothelial cells(EC)are yet to be ultimately determined, we tested variables pertinent to the efficiency of AAV−mediated gene transfer to bovine aortic endothelial cells(BAEC).The variables with AAV vectors were compared with the better characterized adenovirus(Ad)vectors.There is a dose−response relationship between multiplicity of infection(moi)of AAV or Ad vectors and transduction efficiency in BAEC.The higher moi of AAV vectors achieved more than 80% of transduction efficiency in cultured BAEC.AAV and Ad vectors showed an incubation−time−dependent increase in transduction efficiency of LacZ gene to the BAEC up to 12 h of vector exposure.Although the similar kinetics of transduction efficiency of LacZ gene to BAEC was found in both vectors, the duration of gene expression was longer in AAV vector than that in Ad vectors in vitro.These results indicate that AAV−vector is efficient for gene ...
Although adeno-associated virus (AAV)-2 has a broad tissue-host range and can transduce a wide variety of tissue types, some cells, such as erythro-megakaryoblastoid cells, are non-permissive and appear to lack the AAV-2 receptor. However, limited studies have been reported with the related dependovirus AAV-3. We have previously cloned this virus, characterized its genome and produced an infectious clone. In this study, the gene for green fluorescent protein (GFP) was inserted into AAV-2- and AAV-3-based plasmids and recombinant viruses were produced. These viruses were then used to transduce haematopoietic cells and the transduction efficiencies were compared. In contrast to recombinant (r) AAV-2, rAAV-3 successfully transduced erythroid and megakaryoblastoid cells, although rAAV-2 was superior in transduction of lymphocyte-derived cell lines. Recently, it was reported that heparan sulphate can act as a receptor of AAV-2. The infectivity of rAAV-2 and rAAV-3 was tested with mutant cell lines of Chinese
Background- Viral delivery vectors founded on the adeno-associated virus (AAV) serotype 2 platform are currently being evaluated for human gene transfer. All viral genes can be removed, resulting in only the transgene of interest and the viral terminal repeats. These small non-enveloped single-stranded DNA viruses have been associated with long duration expression in experimental animals, and these qualities make AAV serotypes ideal systems when considering viral gene delivery. However, while scores of serotypes have been described, few have been compared directly. In this study we have directly compared AAV serotypes 1-9 by two routes of injection, skeletal muscle and tail vein.. Methods and Results- Previous studies of AAV serotypes 1-5 have demonstrated uniformity in production and purification. Those results guided this comparison of luciferase gene expression-efficiency between AAV serotypes 1-9 after single injection into mouse muscle, or tail vein, of 1 x1011 viral particles into the ...
PURPOSE: Treatment of inherited retinal degenerations using adeno-associated viral (AAV) vectors involves delivery by subretinal injection. In the latter stages, alteration of normal anatomy may cause difficulty in visualizing the retinotomy, retinal detachment extension, and vector diffusion. Vital dyes may be useful surgical adjuncts, but their safety and impact on AAV transduction are largely unknown. METHODS: The effects of Sodium Fluorescein (SF), Membrane Blue (MB), and Membrane Blue Dual (DB) at a range of dilutions were assessed on human embryonic kidney cells in vitro using an AAV2-green fluorescent protein (GFP) reporter at different multiplicities of infection. Flow cytometry analysis was performed to assess both cell viability and transduction efficiency. The effect on quantitative (q)PCR titer was determined. Balanced salt solution (BSS) or dilute DB (1:5 in BSS) were delivered subretinally into left/right eyes of C57BL/6J mice (n = 12). Retinal structure and function were analyzed by
For lentiviral constructs with a fluorescent marker or antibiotic resistance marker, transduction efficiency (i.e., % infected cells) can be determined from the...
ViroTag® AAV5 (for manual sampling) utilizes a fluorescently-labeled, high-affinity antibody which binds to a unique epitope specifically expressed on adeno-associated virus serotype 5. With the Virus Counter 3100, use this rapid, no-wash labeling procedure and take AAV quantification to new levels of accuracy, speed and simplicity!. Product specifications: The ViroTag AAV5 kit (catalog number 92184) contains all reagents and consumables necessary to analyze 200 samples using the Virus Counter 3100 instrument for manual sampling, including:. ...
To assess oncogenic potential, classical transformation assays are based on cell line models. However, cell line based models do not reflect the complexity of human tissues. We thus developed an inducible expression system for gene expression in ex vivo human tissues, which maintain native tissue architecture, such as epithelia and stroma. To validate the system, we transduced and expressed known tumor suppressors (p53, p33ING1b), oncoproteins (RasV12, p47ING3), or controls (empty vector, YFP) in ex vivo prostate tissues, then assessed proliferation by immunohistochemistry of markers (H3S10phos). Herein, we describe how to generate lentiviral vectors and particules, successfully transduce human prostate tissues, induce exogenous gene expression, and assess cellular proliferation.
Problem in stable expression of target protein using lentiviral vector - posted in Cell Biology: To generate cell line to stably express target protein, lenti-viral vector carrying target gene was generated. To validate target gene expression from lentiviral vector, transient transfection was conducted with 293TN cells. Expression of FLAG-tagged target gene was confirmed by Western blot analysis with antibody against FLAG peptide. After generating lentivirus with above lentiviral vector,...
The nucleic acid sequences of adeno-associated virus (AAV) serotype 1 are provided, as are vectors and host cells containing these sequences and functional fragments thereof. Also provided are methods of delivering genes via AAV-1 derived vectors.
ASimilar packaging cells based on the use of T671 cells have been established by the same authors and have been named TeFLY. puro 2 × 105 TU/mL Ref. Rev (transduction by non-SIN MLV vectors) (CMV in the 5LTR tatindependent) SIN, eGFP VSV/G (transient transfection) VSV/G Non-SIN, eGFP MLV ampho, Non-SIN, eGFP GALV, and RD114 Env SIN, eGFP 293 3 × 105 TU/mL 146 293 2 × 105 TU/mL 147 HeLa, 293T HT1080 1 × 107 TU/mL 163 3 × 105 TU/mL 33 With the exception of one report that concerns the SIV LV genus (147), all other publications describe HIV-1-derived packaging cell lines. And Lynch, C. M. (1993) Use of retroviral vectors for gene transfer and expression. Methods Enzymol. 217, 581-599. 15. , et al. (1996) In vivo gene delivery and stable transduction of nondividing cells by a lentiviral vector. Science 272, 263-267. 20 Fluri and Fussenegger 16. Bukovsky, A. , Song, J. , and Naldini, L. (1999) Interaction of human immunodeficiency virus-derived vectors with wild-type virus in transduced cells. ...
AAV-Cre-GFP (AAV Serotype 4) SL101290 AAV4-CMV-Cre-GFP is a pre-packaged rAAV in serotype 4 (with capsid from AAV serotype 4 and 2xITR from AAV serotype 2) whic
Ayuso E, Blouin V, Lock M, McGorray S, Leon X, Alvira MR, Auricchio A, Bucher S, Chtarto A, Clark KR, Darmon C, Doria M, Fountain W, Gao G, Gao K, Giacca M, Kleinschmidt J, Leuchs B, Melas C, Mizukami H, Müller M, Noordman Y, Bockstael O, Ozawa K, Pythoud C, Sumaroka M, Surosky R, Tenenbaum L, van der Linden I, Weins B, Wright JF, Zhang X, Zentilin L, Bosch F, Snyder RO, Moullier P. Manufacturing and characterization of a recombinant adeno-associated virus type 8 reference standard material. Hum Gene Ther. 2014 Nov; 25(11):977-87 ...
ViroTag® AAV2-3 (for auto-sampling) utilizes a fluorescently-labeled, high-affinity antibody which binds to a unique epitope specifically expressed on adeno-associated virus serotypes 2 and 3. With the Virus Counter 3100, use this rapid, no-wash labeling procedure and take AAV quantification to new levels of accuracy, speed and simplicity!. Product specifications: The ViroTag AAV2-3 kit (catalog number 92314) contains all reagents and consumables necessary to analyze 96 samples using the Virus Counter 3100 instrument coupled with an autosampler, including:. ...
Oxford Biomedica used Antha to optimize their lentiviral vector transfection/transduction system and improve production efficiency and robustness. This resulted in ~40 hours of time saving, a 3-10-fold increase in vector titre upon transduction and an 81% reduction in pure error.
Discover Miltenyi Biotecs solutions for isolation, viral transduction, and subsequent phenotypic flow cytometric analysis of hematopoietic stem cells - Latvija
Lentiviral vectors have emerged over the last decade as powerful, reliable and safe tools for stable gene transfer in a wide variety of mammalian cells
Description: The process by which adeno-associated virus (AAV) infects host cells includes viral binding and entry, intracellular trafficking, nuclear transport, and viral second strand DNA synthesis. The second strand DNA synthesis has been shown to be the rate limiting step, which leads to inefficient transduction by AAV vectors. ViraDuctin AAV Transduction Kit is a proprietary, multi-reagent formulation designed to increase the efficiency of transduction by AAV vectors. Reagent preparation takes only 10 minutes prior to host cell infection ...