Pvsv g plasmid

Manufactured by Takara Bio

The PVSV-G plasmid is a commonly used DNA vector that contains the envelope gene of the vesicular stomatitis virus (VSV-G). The VSV-G protein is a viral envelope glycoprotein that facilitates the entry of the virus into host cells. The PVSV-G plasmid is often used in the production of viral vectors, such as lentiviral or retroviral particles, for various research and therapeutic applications.

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Lab products found in correlation

Polybrene, by Merck Group (3 mentions) Pmscvneo vector, by Takara Bio (1 mentions) Pmscvneo, by Takara Bio (1 mentions) Pes filter, by Merck Group (1 mentions) Packaging pcl eco plasmid, by Addgene (1 mentions) Pcmv delta r8.2 plasmid, by Addgene (1 mentions) Lipofectamine 2000 transfection reagent, by Thermo Fisher Scientific (1 mentions) Gp2 293 packaging cell line, by Takara Bio (1 mentions) Lipofectamine 2000 reagent, by Thermo Fisher Scientific (1 mentions)

4 protocols using pvsv g plasmid

Retroviral transduction of myoblasts

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Preparation of retrovirus was performed as reported previously [11 (link)]. Briefly, cDNA for targeting gene (DARP-FLAG) was subcloned into pMSCVneo vector (Clontech). GP2-293 packaging cells were transfected with DARP-pMSCVneo plasmid and pVSV-G plasmid (Clontech) using Lipofectamine 3000. In a parallel way, GP2-293 cells were transfected with GFP-pMSCVneo and pVSV-G plasmids to prepare viruses for negative control. Fresh growth medium was replaced 24 h after transfection, and cells were incubated for another 24h, followed by collection of the virus-containing culture medium. For infection, myoblasts of 50–60% confluency were incubated in the mixture of growth medium and the virus-containing culture medium at 1:1 ratio in the presence of 8 μg/ml polybrene for 24 h. Thereafter, cells were given fresh medium and incubated for 24–48 h followed by protein extraction.

Shimoda Y., Matsuo K., Kitamura Y., Ono K., Ueyama T., Matoba S., Yamada H., Wu T., Chen J., Emoto N, & Ikeda K. (2015). Diabetes-Related Ankyrin Repeat Protein (DARP/Ankrd23) Modifies Glucose Homeostasis by Modulating AMPK Activity in Skeletal Muscle. PLoS ONE, 10(9), e0138624.

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Viral Transduction in HEK 293T Cells

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HEK 293T/17 cells (ATCC® CRL-11268, validated by ATCC) were used no longer than 4 weeks after thawing and were not tested for Mycoplasma. Retroviruses were prepared by co-transfecting HEK 293T/17 cells in a 10-cm plate with 10 μg of packaging pCL-ECO plasmid (Addgene #12371) and 10 μg MIGR1 based vectors by using Lipofectamine 2000 Transfection Reagent (Invitrogen) according to manufacturer’s protocol. Lentiviruses were generated by co-transfection of HEK 293T/17 cells with 10 μg of pLVX-CMV-hTet2(CD)-flag-IRES-mRFP1 based vectors, and 10 μg packaging pCMV delta R8.2 plasmid (Addgene #12263) and envelope pVSV-G plasmid (Clontech, #PT3343-5). Viruses were harvested 40h and 64h after the transfection and filtered through a 0.45-μm PES filter (Millipore). For infection, 5 × 10⁵ cells were suspended in 1 ml of virus-containing medium with 6 μg/ml polybrene (Sigma). GFP+ and RFP+ cells were sorted 48 hrs after the initial infection.

Maifrede S., Le B.V., Nieborowska-Skorska M., Golovine K., Sullivan-Reed K., Dunuwille W.M., Nacson J., Hulse M., Keith K., Madzo J., Caruso L.B., Gazze Z., Lian Z., Padella A., Chitrala K.N., Bartholdy B.A., Matlawska-Wasowska K., Marcantonio D.D., Simonetti G., Greiner G., Sykes S.M., Valent P., Paietta E.M., Tallman M.S., Fernandez H.F., Litzow M.R., Minden M.D., Huang J., Martinelli G., Vassiliou G.S., Tempera I., Piwocka K., Johnson N., Challen G.A, & Skorski T. (2021). TET2 and DNMT3A Mutations Exert Divergent Effects on DNA Repair and Sensitivity of Leukemia Cells to PARP Inhibitors. Cancer research, 81(19), 5089-5101.

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Retroviral Gene Transduction in BOSC23 Cells

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BOSC23 cells were co-transfected with 6 μg of the constructed plasmid plus 1.5 μg of the pVSV-G plasmid (Clontech, Mountain View, CA) using HG TransGene Reagent (Health & Gene, China). After 6 h, the medium was removed and replaced with fresh medium. Viral supernatants were collected, passed through a filter and concentrated. For infection, cells were incubated with serially diluted retroviral supernatants in the presence of 8 μg/ml Polybrene (Sigma, St. Louis, MO), centrifuged at 2,000 rpm for 90 min at 30°C, followed by incubation at 37°C for an additional 6 h. The media was replaced with fresh RPMI 1640 supplemented containing 10% FBS. After 48 h, the cells were treated with SAA for the indicated times, and then harvested for different assays.

Yan Q., Sun L., Zhu Z., Wang L., Li S, & Ye R.D. (2014). Jmjd3-mediated epigenetic regulation of inflammatory cytokine gene expression in serum amyloid A-stimulated macrophages. Cellular signalling, 26(9), 1783-1791.

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Genotyping and Overexpression of Ovarian Cancer Antigens

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HLA DPB1 genotype for ovarian cancer cell lines was determined by the SSP-PCR method39 (link). Protein-coding regions of NY-ESO-1 and Sp17 were PCR amplified from cDNA of SK37 and cloned into the first multiple cloning site in a pQCXIX self-inactivating retroviral vector (Clontech). CIITA-coding DNA was also amplified from SK37 cDNA and cloned into the second multiple cloning site in the same vector following the internal ribosomal entry site (IRES). Resulting pQC-NY-ESO-1-IRES-CIITA or pQC-Sp17-IRES-CIITA plasmid was co-transfected together with a pVSV-G plasmid (Clontech) into a GP2-293 packaging cell line (Clontech) using Lipofectamine 2000 reagent (Invitrogen). Retroviral supernatant was added on DP4⁺ ovarian cancer cell lines (OV2774 and OVCAR-5) in the presence of 8 μg/ml polybrene (Sigma). Five to 10 days after infection, cells were stained with anti-HLA-DR antibody and HLA-DR⁺ cells were sorted by a FACSAria instrument.

Matsuzaki J., Tsuji T., Luescher I.F., Shiku H., Mineno J., Okamoto S., Old L.J., Shrikant P., Gnjatic S, & Odunsi K. (2015). Direct tumor recognition by a human CD4+ T-cell subset potently mediates tumor growth inhibition and orchestrates anti-tumor immune responses. Scientific Reports, 5, 14896.

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