Plenti c mgfp

Manufactured by OriGene

Sourced in United States

The PLenti-C-mGFP is a lentiviral vector that expresses monomeric green fluorescent protein (mGFP) under the constitutive CMV promoter. It is designed for the stable transduction of cells and expression of the mGFP reporter gene.

Automatically generated - may contain errors

Lab products found in correlation

Lenticrisprv2 blast plasmid, by Addgene (2 mentions) N174 mcs, by Addgene (2 mentions) Plenti cd1d1 mgfp, by OriGene (2 mentions) Plenti ifi204 myc ddk puro, by OriGene (2 mentions) Fastdigest protocol, by Thermo Fisher Scientific (2 mentions) Nebuilder hifi dna assembly cloning kit, by New England Biolabs (2 mentions) Pdonr223 axl, by Addgene (1 mentions) Facsaria 3, by BD (1 mentions) Stereotaxic frame, by Stoelting (1 mentions) Plenticrisprv2, by Addgene (1 mentions) Lipofectamine 3000 system, by Thermo Fisher Scientific (1 mentions) Eclipse e600 microscope, by Nikon (1 mentions) Prolong gold antifade mountant with dapi, by Thermo Fisher Scientific (1 mentions) Image pro software, by Media Cybernetics (1 mentions) Quickchange primer design, by Agilent Technologies (1 mentions) Pgfp c shlenti vector, by OriGene (1 mentions) Td 20 20 luminometer, by Turner Designs (1 mentions) Dual luciferase assay kit, by Promega (1 mentions) Lipfectamine 2000, by Thermo Fisher Scientific (1 mentions) In fusion enzyme, by Takara Bio (1 mentions)

15 protocols using plenti c mgfp

Overexpression of PKCδ Constructs in Cancer Cells

Check if the same lab product or an alternative is used in the 5 most similar protocols

PKCδ constructs were purchased from Addgene (75 Sidney Street, Suite 550A Cambridge, MA 02139). PKCδ-C2 domain (Addgene #16388) and wild-type PKCδ (Addgene #16386) were cloned into lentiviral vector pLenti-C-mGFP from OriGene (9620 Medical Center Dr., Suite 200, Rockville, MD 20850) (PS100071) via EcoRI digestion and ligation and transfected via electroporation using a Neon transfection system. 48 h later, positive cells were selected by FACS sorting GFP + population.
PKCδ DN was a gift from Bernard Weinstein (Addgene plasmid # 16389). PKCδ DN contains a K376R mutation. PKC8 DN and wild-type PKCδ (Plasmid #16386) were cloned into OriGene pLenti-C-mGFP (PS100071) and transfected via electroporation into E⁻DU145 shCDCP1. After 48 h, cells positive for PKC8 DN were selected via FACS sorting the GFP + population.

Pollan S.G., Huang F., Sperger J.M., Lang J.M., Morrissey C., Cress A.E., Chu C.Y., Bhowmick N.A., You S., Freeman M.R., Spassov D.S., Moasser M.M., Carter W.G., Satapathy S.R., Shah K, & Knudsen B.S. (2018). Regulation of inside-out β1-integrin activation by CDCP1. Oncogene, 37(21), 2817-2836.

+ Open protocol

+ Expand

Wnt Pathway Modulation in 293FT and 3T3 Cells

Check if the same lab product or an alternative is used in the 5 most similar protocols

We transfected 293FT cells grown to 80% confluency in a 12-well plate, using Lipfectamine 2000 (Invitrogen, Carlsbad, CA), with 500 ng of DNA. All cells were transfected with TOPflash or FOPflash reporter plasmids alongside a vector force-expressing ARID3B (pLenti-suCMV-Rsv, Gentarget, San Diego, CA), FZD5 (pLenti-C-mGFP, Origene, Rockville, MD) Wnt5a (pLenti-C-mGFP), or an empty pLenti-C-mGFP vector. All samples were also cotransfected with a Renilla luciferase control to normalize for cell number and transfection efficiency. Luciferase activity was measured using a Promega Dual Luciferase Assay Kit (Promega, Madison, WI) according to the manufacturer's instructions and measured on a TD 20/20 luminometer (Turner Designs, Sunnyvale, CA). A similar experiment was conducted with 3T3 "Leading Light" cells (Enzo Life Sciences, Farmingdale, NY). As these cells express a Wnt Reporter luciferase, they were only transfected with the above-mentioned ARID3B, FZD5, and Wnt5a vectors, and normalized by protein concentration. All conditions were run in triplicate and normalized to the Renilla internal control.

Bobbs A., Gellerman K., Hallas W.M., Joseph S., Yang C., Kurkewich J, & Cowden Dahl K.D. (2015). ARID3B Directly Regulates Ovarian Cancer Promoting Genes. PLoS ONE, 10(6), e0131961.

+ Open protocol

+ Expand

Cloning and Overexpression of AXL

Cited 1 time

Check if the same lab product or an alternative is used in the 5 most similar protocols

pDONR223-AXL was obtained from Addgene, Cambridge, MA, USA (Addgene plasmid # 23945) [33 (link)]. AXL cDNA was cloned into the pLenti-C-mGFP (OriGene, Rockville, MD, USA) and SCC-25 cells were transduced with this vector or empty vector as control using lentiviral transduction. Fluorescence-activated cell sorting (FACS) of GFP+ cells (FACSAriaIII, BD Biosciences, Heidelberg, Germany) was used to select positive cells. Overexpression was subsequently estimated via Western blot.

von Mässenhausen A., Brägelmann J., Billig H., Thewes B., Queisser A., Vogel W., Kristiansen G., Schröck A., Bootz F., Brossart P., Kirfel J, & Perner S. (2016). Implication of the Receptor Tyrosine Kinase AXL in Head and Neck Cancer Progression. International Journal of Molecular Sciences, 18(1), 7.

+ Open protocol

+ Expand

Lentiviral Overexpression of Cyclophilin D in Mice

Check if the same lab product or an alternative is used in the 5 most similar protocols

Animals received bilateral intrahippocampal administration of Lenti ORF particles (GFP-tagged)-mouse peptidylprolyl isomerase D (Ppid; cyclophilin D) (Type: Human Tagged ORF Clone Lentiviral Particle; Tag: mGFP; Vector: pLenti-C-mGFP, #RC223397L2V, Origene) by stereotaxic injection (Vargas et al., 2014 (link)). A total of n = 5 mice were used per group. We used Sham injection in WT and control tau KO mice. Tau−/− mice were anesthetized using isoflurane and placed in a stereotaxic frame (Stoelting, United States). The skull was exposed for several millimeters anterior and posterior to the bregma. Boreholes were made above the left and right hippocampal CA1 (coordinates: 2.46 mm anterior to the bregma, 1.0 mm lateral, 1.5 mm relative to dura mater). One microliter of the lentiviral vector was injected (10⁸ TU/ml). Three weeks after infection, the animals were subjected to cognitive tests and euthanized immediately after for biochemical analysis.

Jara C., Cerpa W., Tapia-Rojas C, & Quintanilla R.A. (2021). Tau Deletion Prevents Cognitive Impairment and Mitochondrial Dysfunction Age Associated by a Mechanism Dependent on Cyclophilin-D. Frontiers in Neuroscience, 14, 586710.

+ Open protocol

+ Expand

CRISPR Gene Disruption and Overexpression Validation

Check if the same lab product or an alternative is used in the 5 most similar protocols

To validate the candidate genes from screening, sgRNAs from the parent library were cloned into pLentiCRISPRv2 (Addgene plasmid 52961). The control sgRNA was used from the parent library. Lentiviruses were produced as described above, and transduced HAP1 or HeLa cells were selected with 1 μg/ml puromycin 24 h post-infection. Two weeks later, puromycin was removed, and cells were allowed to recover for three additional days before analysis. Gene disruption efficiency was verified by Western blot. The sequences of the sgRNAs used are in Additional file 1: Table S9.
The gRNA-resistant construct (OE_RDD1) was made by directed mutagenesis using the Quick-Change kit (Stratagene) following the manufacturer’s protocol. To create the OE_RDD1 construct, the sgRNA targeting site (sgRDD1_B; Additional file 1: Table S9) on pLenti-C-mGFP (Origene) containing human C1orf115 gene (pLenti-C1orf115-mGFP) was changed to gRNA resistance sequence (synonymous mutations) using the following oligonucleotide—sense: 5′-GCC GCT TAT AGC GCT CCT TTC GCT GTA GCC ACC AGC GTG GTA TCC-3′, and anti-sense: 5′-AGC GAA AGG AGC GCT ATA AGC GGC AGC GAA GCC TTG CAG GCC G-3′.

Lau M.T., Ghazanfar S., Parkin A., Chou A., Rouaen J.R., Littleboy J.B., Nessem D., Khuong T.M., Nevoltris D., Schofield P., Langley D., Christ D., Yang J., Pajic M, & Neely G.G. (2020). Systematic functional identification of cancer multi-drug resistance genes. Genome Biology, 21, 27.

+ Open protocol

+ Expand

TGFBI Overexpression in HPAECs

Check if the same lab product or an alternative is used in the 5 most similar protocols

HPAECs were transduced with lentiviral particles (OriGene; control particles pLenti-C-mGFP, PS100071; or TGFBI Lentiviral Particles, RC200411L2V), following the instructions of the manufacturer. Forty-eight hours later, cells were either fixed in 4% paraformaldehyde for immunofluorescence staining with primary antibody (polyclonal TGFBI antibody ab99562; Abcam) or lysed in Trizol for mRNA isolation.

Bochenek M.L., Saar K., Nazari-Jahantigh M., Gogiraju R., Wiedenroth C.B., Münzel T., Mayer E., Fink L., Schober A., Hübner N., Guth S., Konstantinides S, & Schäfer K. (2023). Endothelial Overexpression of TGF-β-Induced Protein Impairs Venous Thrombus Resolution: Possible Role in CTEPH. JACC: Basic to Translational Science, 9(1), 100-116.

+ Open protocol

+ Expand

Transient Transfection and Transduction Efficiency Assay

Check if the same lab product or an alternative is used in the 5 most similar protocols

The plasmid constructs used for transient transfection and transduction were pXOON-eGFP (Addgene, Cambridge MA) and pLenti-C-mGFP (OriGene, Rockville MD), respectively. ACC cells were plated on cover slips in 24-well plate for 24 hrs prior to transient transfection and transduction. For transfection we used Lipofectamine 3000 system (ThermoFisher Scientific, Carlsbad CA) per manufacture’s protocol. Transient transduction using pLenti-C-mGFP was performed according to the standard protocol, and cell were fixed 48% after transduction. Stable cell lines were generated according to the standard protocol with plasmid selection. Cells were grown on coverslips for 48hr, and fixed using ProLong Gold Antifade Mountant with DAPI (ThermoFisher Scientific, Carlsbad CA). Slides were imaged using Nikon Eclipse E600 microscope (Nikon, Tokyo, Japan) equipped with a CoolSnap fx camera (Photometrics, Tucson, AZ, USA) and using ImagePro software (Media Cybernetics, Rockville, MD, USA). Cells that were positive for GFP were directly counted and normalized against the number of cells that were positive for DAPI in any given field. Five such fields were counted for each slide per experiment and average of three experimental replicates was used to calculate the transient transfection and transduction efficiency.

Kiseljak-Vassiliades K., Zhang Y., Bagby S.M., Kar A., Pozdeyev N., Xu M., Gowan K., Sharma V., Raeburn C.D., Albuja-Cruz M., Jones K.L., Fishbein L., Schweppe R.E., Somerset H., Pitts T.M., Leong S, & Wierman M.E. (2018). Development of New Preclinical Models to Advance Adrenocortical Carcinoma Research. Endocrine-related cancer, 25(4), 437-451.

+ Open protocol

+ Expand

Lentiviral Transduction and Migration Assay

Check if the same lab product or an alternative is used in the 5 most similar protocols

Plasmids such as pLenti-C-mGFP and pLenti-C-mGFP-KLF6 were purchased from Origene. Virus particles were packaged according to the manufacturer's recommendation and infected into SAS cells. Twenty-four hour post-infection, cells were seeded at a density of 5 × 105/mL in the upper chamber of the 48-well Boyden chamber. The lower chamber contained 20% FBS. The chamber was incubated at 37 °C for 24 h. The cells that migrated to the lower surface of the membrane were fixed in methanol for 10 min and stained with 10% Giemsa for 1 h. The migrated cells were pictured at random five fields and counted.

Hsu L.S., Huang R.H., Lai H.W., Hsu H.T., Sung W.W., Hsieh M.J., Wu C.Y., Lin Y.M., Chen M.K., Lo Y.S, & Chen C.J. (2017). KLF6 inhibited oral cancer migration and invasion via downregulation of mesenchymal markers and inhibition of MMP-9 activities. International Journal of Medical Sciences, 14(6), 530-535.

+ Open protocol

+ Expand

Overexpression and Knockdown of PRMT7

Check if the same lab product or an alternative is used in the 5 most similar protocols

To overexpress either C-terminal Myc-tagged or GFP-tagged PRMT7, pLenti-C-MycDDK or pLenti-C-mGFP, respectively, were purchased from Origene. A control vector expressing only GFP or Myc was created via blunt-end digestion and subsequent ligation of the pLenti-C-mGFP-PRMT7 and pLenti-C-MycDDK-PRMT7 vectors, respectively (characterized in Baldwin et al., 2015 (link)). Mutant eIF2α-GST protein from pGEX-4T2 vector was designed: R⁵²R⁵³R⁵⁴ mutated to KKK, RKK, KRR, RKR, and RRK. Primers were designed using QuickChange Primer Design by Agilent Technologies. These constructs were then subcloned into the pLenti-C-mGFP backbone as described above. To knock down PRMT7, RNA interference was performed using pLKO.1 vectors obtained from The RNAi Consortium containing either an shRNA with a luciferase sequence for control (5′-CAAATCACAGAATCGTCGTAT-3′) or two independent PRMT7 sequences (5′-GCTAACCACTTGGAAGATAAA-3′ and 5′-CGATGACTACTGCGTATGGTA-3′).

Haghandish N., Baldwin R.M., Morettin A., Dawit H.T., Adhikary H., Masson J.Y., Mazroui R., Trinkle-Mulcahy L, & Côté J. (2019). PRMT7 methylates eukaryotic translation initiation factor 2α and regulates its role in stress granule formation. Molecular Biology of the Cell, 30(6), 778-793.

+ Open protocol

+ Expand

Lentiviral CD1d1 and IFI204 constructs

Check if the same lab product or an alternative is used in the 5 most similar protocols

A lentiviral murine Cd1d1 expression construct (pLenti-Cd1d1-mGFP, Cat#MR226027L4) and its matched control construct (pLenti-C-mGFP, Cat#PS100093) were obtained from Origene. Murine Ifi204 expression vector (pLenti-Ifi204-Myc-DDK-Puro, Cat#MR222527L3), together with its control vector (pLenti-C-Myc-DDK-Puro, Cat#PS100092) were also purchased from Origene, and the puromycin selection cassette of these two Origene plasmids were replaced by blasticidin from lentiCRISPRv2-blast plasmid (Addgene#98293) using NEBuilder HiFi DNA Assembly Cloning kit (NEB, Cat#E5520S). For the LLC-1 experiment, the murine Cd1d1 was cloned into the receiving vector N174-MCS (Addgene#81061) with the restriction enzymes EcoR1 and Mlu1, using the FastDigest protocol of Thermo Scientific. All final construct sequences were confirmed by Sanger sequencing. Plasmids generated in this study are available upon written request.

Guo H., Vuille J.A., Wittner B.S., Lachtara E.M., Hou Y., Lin M., Zhao T., Raman A.T., Russell H.C., Reeves B.A., Pleskow H.M., Wu C.L., Gnirke A., Meissner A., Efstathiou J.A., Lee R.J., Toner M., Aryee M.J., Lawrence M.S., Miyamoto D.T., Maheswaran S, & Haber D.A. (2023). DNA hypomethylation silences antitumor immune genes in early prostate cancer and CTCs. Cell, 186(13), 2765-2782.e28.

+ Open protocol

+ Expand

About PubCompare

Our mission is to provide scientists with the largest repository of trustworthy protocols and intelligent analytical tools, thereby offering them extensive information to design robust protocols aimed at minimizing the risk of failures.

We believe that the most crucial aspect is to grant scientists access to a wide range of reliable sources and new useful tools that surpass human capabilities.

However, we trust in allowing scientists to determine how to construct their own protocols based on this information, as they are the experts in their field.

Ready to get started?

Revolutionizing how scientists
search and build protocols!