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third generation lentiviral packaging plasmids pmdlg prre  (Addgene inc)


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    Structured Review

    Addgene inc third generation lentiviral packaging plasmids pmdlg prre
    Loss of BCAT1 impacts hiPSC-derived cortical neuron differentiation (A) Schematic of hiPSC <t>lentiviral</t> transduction for Ngn2-iN and experimental design. (B) Representative western blot analysis of BCAT1 E/K F/L and BCAT1 −/− Ngn2-iNs collected at day 2 and day 14 of differentiation (left). Quantification of vinculin expression in day 14 neurons (right). Results are mean ± SEM ( n = 3 independent experiments per group in all 3 clonal lines) and analyzed by one-way ANOVA with Dunnett’s post hoc test. (C) qRT-PCR analysis of BCAT1 , BCAT2 , and NeuroD1 transcript levels in day 14 Ngn2-iNs normalized to RPLP0 . Results represented as means ± SEM ( n ≥ 3 independent experiments per group in all 3 clonal lines), analyzed by two-way ANOVA with Dunnett’s post hoc analysis test for multiple comparisons. (D) Representative confocal images and (E) fluorescence intensity measurements of developing neurons 2, 7, and 14 days post-differentiation (dpd) showing increased Tuj1 and Map2 expression. Scale bars, 100 μm. Immunocytochemistry was performed in all three clonal lines and two cell culture replicates per time point, with similar results. (F) Reconstructions of sparse-labeled Ngn2-iNs derived from BCAT1 E/K F/L and isogenic control lines. Experiments were performed in all two cell culture replicates per time point. Results are shown in (G). BCAT1 neurons exhibit reduced growth properties and were significantly shorter at 2 dpd (left) compared with 7 dpd (right) (two-way ANOVA with Sidak correction; 2 dpd: ∗∗ p < 0.005; 7 dpi: n.s., not significant). Values show means ± SEM.; BCAT1 E/K F/L ( n = 1; 33 technical tracing replicates in total); control ( n = 1; 29 technical tracing replicates in total); n refers to biologically independent clonal lines. A.U., arbitrary units.
    Third Generation Lentiviral Packaging Plasmids Pmdlg Prre, supplied by Addgene inc, used in various techniques. Bioz Stars score: 96/100, based on 1625 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/product/third+generation+lentiviral+packaging+system/pMDLg%2FpRRE+(Plasmid+%2312251)/pmc12670911-83-18-23
    Average 96 stars, based on 1625 article reviews
    third generation lentiviral packaging plasmids pmdlg prre - by Bioz Stars, 2026-09
    96/100 stars

    Images

    1) Product Images from "Bi-allelic variants in BCAT1 impair mitochondrial function and are associated with a candidate neurometabolic disorder"

    Article Title: Bi-allelic variants in BCAT1 impair mitochondrial function and are associated with a candidate neurometabolic disorder

    Journal: Human Genetics and Genomics Advances

    doi: 10.1016/j.xhgg.2025.100525

    Loss of BCAT1 impacts hiPSC-derived cortical neuron differentiation (A) Schematic of hiPSC lentiviral transduction for Ngn2-iN and experimental design. (B) Representative western blot analysis of BCAT1 E/K F/L and BCAT1 −/− Ngn2-iNs collected at day 2 and day 14 of differentiation (left). Quantification of vinculin expression in day 14 neurons (right). Results are mean ± SEM ( n = 3 independent experiments per group in all 3 clonal lines) and analyzed by one-way ANOVA with Dunnett’s post hoc test. (C) qRT-PCR analysis of BCAT1 , BCAT2 , and NeuroD1 transcript levels in day 14 Ngn2-iNs normalized to RPLP0 . Results represented as means ± SEM ( n ≥ 3 independent experiments per group in all 3 clonal lines), analyzed by two-way ANOVA with Dunnett’s post hoc analysis test for multiple comparisons. (D) Representative confocal images and (E) fluorescence intensity measurements of developing neurons 2, 7, and 14 days post-differentiation (dpd) showing increased Tuj1 and Map2 expression. Scale bars, 100 μm. Immunocytochemistry was performed in all three clonal lines and two cell culture replicates per time point, with similar results. (F) Reconstructions of sparse-labeled Ngn2-iNs derived from BCAT1 E/K F/L and isogenic control lines. Experiments were performed in all two cell culture replicates per time point. Results are shown in (G). BCAT1 neurons exhibit reduced growth properties and were significantly shorter at 2 dpd (left) compared with 7 dpd (right) (two-way ANOVA with Sidak correction; 2 dpd: ∗∗ p < 0.005; 7 dpi: n.s., not significant). Values show means ± SEM.; BCAT1 E/K F/L ( n = 1; 33 technical tracing replicates in total); control ( n = 1; 29 technical tracing replicates in total); n refers to biologically independent clonal lines. A.U., arbitrary units.
    Figure Legend Snippet: Loss of BCAT1 impacts hiPSC-derived cortical neuron differentiation (A) Schematic of hiPSC lentiviral transduction for Ngn2-iN and experimental design. (B) Representative western blot analysis of BCAT1 E/K F/L and BCAT1 −/− Ngn2-iNs collected at day 2 and day 14 of differentiation (left). Quantification of vinculin expression in day 14 neurons (right). Results are mean ± SEM ( n = 3 independent experiments per group in all 3 clonal lines) and analyzed by one-way ANOVA with Dunnett’s post hoc test. (C) qRT-PCR analysis of BCAT1 , BCAT2 , and NeuroD1 transcript levels in day 14 Ngn2-iNs normalized to RPLP0 . Results represented as means ± SEM ( n ≥ 3 independent experiments per group in all 3 clonal lines), analyzed by two-way ANOVA with Dunnett’s post hoc analysis test for multiple comparisons. (D) Representative confocal images and (E) fluorescence intensity measurements of developing neurons 2, 7, and 14 days post-differentiation (dpd) showing increased Tuj1 and Map2 expression. Scale bars, 100 μm. Immunocytochemistry was performed in all three clonal lines and two cell culture replicates per time point, with similar results. (F) Reconstructions of sparse-labeled Ngn2-iNs derived from BCAT1 E/K F/L and isogenic control lines. Experiments were performed in all two cell culture replicates per time point. Results are shown in (G). BCAT1 neurons exhibit reduced growth properties and were significantly shorter at 2 dpd (left) compared with 7 dpd (right) (two-way ANOVA with Sidak correction; 2 dpd: ∗∗ p < 0.005; 7 dpi: n.s., not significant). Values show means ± SEM.; BCAT1 E/K F/L ( n = 1; 33 technical tracing replicates in total); control ( n = 1; 29 technical tracing replicates in total); n refers to biologically independent clonal lines. A.U., arbitrary units.

    Techniques Used: Derivative Assay, Transduction, Western Blot, Expressing, Quantitative RT-PCR, Fluorescence, Immunocytochemistry, Cell Culture, Labeling, Control

    Related Articles

    Construct:

    Article Title: Development of a Synthetic 3-ketosteroid Δ 1 -dehydrogenase for the Generation of a Novel Catabolic Pathway Enabling Cholesterol Degradation in Human Cells.
    Article Snippet: .. Lentiviral particles encoding the Δ1-KstDR constructs were produced with HEK293FT cells using the third-generation lentiviral packaging system (Addgene #s: 12251, 12253, and 12259). .. Packaging vectors (7.5 μg pMDLg/pRRE, 3.75 μg RSV-REV, and 4.5 μg PMD2.G) and each transfer vector (3 μg pLenti-CMV-Blast (706-1)-Δ1-KstDR, pLenti-CMV-Puro (w118-1)-Δ1-KstDR, or pLenti-PGK-Hygro-(w530-1)Δ1-KstDR) were diluted with 1.875 mL Opti-MEM (1 μg plasmid DNA/100 μL Opti-MEM) in a glass vial.

    Article Title: PMS2 variant results in loss of ATPase activity without compromising mismatch repair
    Article Snippet: The 2653 bp amplicons were agarose gel purified (Qiagen), cloned into pDonr221 using Gateway recombination, further subcloned into pLenti‐CMV‐Blast‐Dest (706–1) (Campeau et al., ), and sequence verified. .. Lentiviral particles encoding PMS2 WT, p.Asn335Ser, and p.Leu42_Glu44del constructs were produced with HEK‐293FT cells using the third‐generation lentiviral packaging system (Addgene # 12251, 12,253, and 12,259). .. Each transfer vector (3 μg pLenti‐CMV‐Blast (706–1)‐PMS2 WT, p.Asn335Ser, and p.Leu42_Glu44del) and packaging vectors (7.5 μg pMDLg/pRRE, 3.75 μg RSV‐REV, and 4.5 μg PMD2.G) were diluted with 1.875 ml Opti‐MEM (1 μg plasmid DNA/100 μl Opti‐MEM) in a glass vial.

    Produced:

    Article Title: Development of a Synthetic 3-ketosteroid Δ 1 -dehydrogenase for the Generation of a Novel Catabolic Pathway Enabling Cholesterol Degradation in Human Cells.
    Article Snippet: .. Lentiviral particles encoding the Δ1-KstDR constructs were produced with HEK293FT cells using the third-generation lentiviral packaging system (Addgene #s: 12251, 12253, and 12259). .. Packaging vectors (7.5 μg pMDLg/pRRE, 3.75 μg RSV-REV, and 4.5 μg PMD2.G) and each transfer vector (3 μg pLenti-CMV-Blast (706-1)-Δ1-KstDR, pLenti-CMV-Puro (w118-1)-Δ1-KstDR, or pLenti-PGK-Hygro-(w530-1)Δ1-KstDR) were diluted with 1.875 mL Opti-MEM (1 μg plasmid DNA/100 μL Opti-MEM) in a glass vial.

    Article Title: PMS2 variant results in loss of ATPase activity without compromising mismatch repair
    Article Snippet: The 2653 bp amplicons were agarose gel purified (Qiagen), cloned into pDonr221 using Gateway recombination, further subcloned into pLenti‐CMV‐Blast‐Dest (706–1) (Campeau et al., ), and sequence verified. .. Lentiviral particles encoding PMS2 WT, p.Asn335Ser, and p.Leu42_Glu44del constructs were produced with HEK‐293FT cells using the third‐generation lentiviral packaging system (Addgene # 12251, 12,253, and 12,259). .. Each transfer vector (3 μg pLenti‐CMV‐Blast (706–1)‐PMS2 WT, p.Asn335Ser, and p.Leu42_Glu44del) and packaging vectors (7.5 μg pMDLg/pRRE, 3.75 μg RSV‐REV, and 4.5 μg PMD2.G) were diluted with 1.875 ml Opti‐MEM (1 μg plasmid DNA/100 μl Opti‐MEM) in a glass vial.

    shRNA:

    Article Title: De Novo Multi-Omics Pathway Analysis Designed for Prior Data Independent Inference of Cell Signaling Pathways.
    Article Snippet: In Brief A novel tool, de novo multiomics pathway analysis (DMPA), is introduced for prior data independent and unbiased cell signaling pathway inference from multi-omics data.. DMPA was validated with available omics data and outperformed previous module and pathway inference methods, especially with datasets of low sample sizes.. DMPA includes upstream regulator, subcellular location, and biological function prediction tools for modules and pathways.

    Article Title: De Novo Multi-Omics Pathway Analysis Designed for Prior Data Independent Inference of Cell Signaling Pathways
    Article Snippet: The control lentiviral shRNA plasmid (Addgene plasmid #1864) was a kind gift from David Sabatini ( ). .. Third generation lentiviral packaging system (Addgene) was used in HEK293T cells to produce shRNA-carrying lentiviruses. ..

    Article Title: Resveratrol derivative SH-707 inhibits NLRP3 inflammasome activation via a sirtuin 1-dependent pathway.
    Article Snippet: Lentivirus was produced by transfecting HEK293T cells (CRL11268; ATCC, Manassas, VA, USA). .. The cells were seeded in 6-well plates, and the lentivirus was assembled using a third-generation lentiviral packaging system (pMDLg/pRRE, #12251; pRSV-Rev, #12253; pMD2.G, #12259; and Sirt1 [sc-40987-SH] shRNA plasmid purchased from Addgene and Santa Cruz, MISSION® pLKO.1-puro Non-Target shRNA Control Plasmid DNA [SHC016] purchased from Sigma-Aldrich). ..

    Article Title: An unbiased pathway analysis (UPA) designed for multi-omics inference of cell signaling pathways
    Article Snippet: The control lentiviral shRNA plasmid (Addgene plasmid #1864) was a kind gift from David Sabatini ( Sarbassov et al , 2005 ). .. Third generation lentiviral packaging system (Addgene) was used in HEK293T cells to produce shRNA-carrying lentiviruses. ..

    Knockdown:

    Article Title: ANO1 Reprograms Cholesterol Metabolism and the Tumor Microenvironment to Promote Cancer Metastasis
    Article Snippet: All plasmids used in this study were subcloned to corresponding vectors by homologous recombination method using pEASY-Basic Seamless Cloning and Assembly Kit (Transgen). .. ANO1-overexpressing or -knockdown ESCC cell lines were established using third-generation lentiviral packaging system (Addgene#12251, Addgene12253, Addgene#12259) and maintained with Puromycin (Thermo Fisher Scientific). ..

    Plasmid Preparation:

    Article Title: Resveratrol derivative SH-707 inhibits NLRP3 inflammasome activation via a sirtuin 1-dependent pathway.
    Article Snippet: Lentivirus was produced by transfecting HEK293T cells (CRL11268; ATCC, Manassas, VA, USA). .. The cells were seeded in 6-well plates, and the lentivirus was assembled using a third-generation lentiviral packaging system (pMDLg/pRRE, #12251; pRSV-Rev, #12253; pMD2.G, #12259; and Sirt1 [sc-40987-SH] shRNA plasmid purchased from Addgene and Santa Cruz, MISSION® pLKO.1-puro Non-Target shRNA Control Plasmid DNA [SHC016] purchased from Sigma-Aldrich). ..

    Control:

    Article Title: Resveratrol derivative SH-707 inhibits NLRP3 inflammasome activation via a sirtuin 1-dependent pathway.
    Article Snippet: Lentivirus was produced by transfecting HEK293T cells (CRL11268; ATCC, Manassas, VA, USA). .. The cells were seeded in 6-well plates, and the lentivirus was assembled using a third-generation lentiviral packaging system (pMDLg/pRRE, #12251; pRSV-Rev, #12253; pMD2.G, #12259; and Sirt1 [sc-40987-SH] shRNA plasmid purchased from Addgene and Santa Cruz, MISSION® pLKO.1-puro Non-Target shRNA Control Plasmid DNA [SHC016] purchased from Sigma-Aldrich). ..

    Transfection:

    Article Title: The inflammasome-activating poxvirus peptide IAMP29 promotes antimicrobial and anticancer responses
    Article Snippet: Lentiviral shRNA preparation and knockdown of NLRP3 and PKM2 Lentiviruses were produced by transfecting HEK293T cells (ATCC, Manassas, VA, USA; CRL-11268) seeded the previous day in 6-well plates using a three-plasmid system. .. When the HEK293T cells reached 60–70% confluency, they were transfected with pLV[shRNA]-Puro-U6 plasmids targeting hNLRP3 (5’-GCGTTAGAAACACTTCAAGAA-3’) and hPKM2 (5’-AGGCAGAGGCTGCCATCTACC-3’) (VectorBuilder, Chicago, IL, USA), along with a third-generation lentiviral packaging system (pRSV-Rev, #12253; pMD2.G, #12259; and pMDLg/pRRE, #12251, obtained from Addgene). ..



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    Image Search Results


    Loss of BCAT1 impacts hiPSC-derived cortical neuron differentiation (A) Schematic of hiPSC lentiviral transduction for Ngn2-iN and experimental design. (B) Representative western blot analysis of BCAT1 E/K F/L and BCAT1 −/− Ngn2-iNs collected at day 2 and day 14 of differentiation (left). Quantification of vinculin expression in day 14 neurons (right). Results are mean ± SEM ( n = 3 independent experiments per group in all 3 clonal lines) and analyzed by one-way ANOVA with Dunnett’s post hoc test. (C) qRT-PCR analysis of BCAT1 , BCAT2 , and NeuroD1 transcript levels in day 14 Ngn2-iNs normalized to RPLP0 . Results represented as means ± SEM ( n ≥ 3 independent experiments per group in all 3 clonal lines), analyzed by two-way ANOVA with Dunnett’s post hoc analysis test for multiple comparisons. (D) Representative confocal images and (E) fluorescence intensity measurements of developing neurons 2, 7, and 14 days post-differentiation (dpd) showing increased Tuj1 and Map2 expression. Scale bars, 100 μm. Immunocytochemistry was performed in all three clonal lines and two cell culture replicates per time point, with similar results. (F) Reconstructions of sparse-labeled Ngn2-iNs derived from BCAT1 E/K F/L and isogenic control lines. Experiments were performed in all two cell culture replicates per time point. Results are shown in (G). BCAT1 neurons exhibit reduced growth properties and were significantly shorter at 2 dpd (left) compared with 7 dpd (right) (two-way ANOVA with Sidak correction; 2 dpd: ∗∗ p < 0.005; 7 dpi: n.s., not significant). Values show means ± SEM.; BCAT1 E/K F/L ( n = 1; 33 technical tracing replicates in total); control ( n = 1; 29 technical tracing replicates in total); n refers to biologically independent clonal lines. A.U., arbitrary units.

    Journal: Human Genetics and Genomics Advances

    Article Title: Bi-allelic variants in BCAT1 impair mitochondrial function and are associated with a candidate neurometabolic disorder

    doi: 10.1016/j.xhgg.2025.100525

    Figure Lengend Snippet: Loss of BCAT1 impacts hiPSC-derived cortical neuron differentiation (A) Schematic of hiPSC lentiviral transduction for Ngn2-iN and experimental design. (B) Representative western blot analysis of BCAT1 E/K F/L and BCAT1 −/− Ngn2-iNs collected at day 2 and day 14 of differentiation (left). Quantification of vinculin expression in day 14 neurons (right). Results are mean ± SEM ( n = 3 independent experiments per group in all 3 clonal lines) and analyzed by one-way ANOVA with Dunnett’s post hoc test. (C) qRT-PCR analysis of BCAT1 , BCAT2 , and NeuroD1 transcript levels in day 14 Ngn2-iNs normalized to RPLP0 . Results represented as means ± SEM ( n ≥ 3 independent experiments per group in all 3 clonal lines), analyzed by two-way ANOVA with Dunnett’s post hoc analysis test for multiple comparisons. (D) Representative confocal images and (E) fluorescence intensity measurements of developing neurons 2, 7, and 14 days post-differentiation (dpd) showing increased Tuj1 and Map2 expression. Scale bars, 100 μm. Immunocytochemistry was performed in all three clonal lines and two cell culture replicates per time point, with similar results. (F) Reconstructions of sparse-labeled Ngn2-iNs derived from BCAT1 E/K F/L and isogenic control lines. Experiments were performed in all two cell culture replicates per time point. Results are shown in (G). BCAT1 neurons exhibit reduced growth properties and were significantly shorter at 2 dpd (left) compared with 7 dpd (right) (two-way ANOVA with Sidak correction; 2 dpd: ∗∗ p < 0.005; 7 dpi: n.s., not significant). Values show means ± SEM.; BCAT1 E/K F/L ( n = 1; 33 technical tracing replicates in total); control ( n = 1; 29 technical tracing replicates in total); n refers to biologically independent clonal lines. A.U., arbitrary units.

    Article Snippet: Ngn2-iN lentiviruses were produced as previously described in HEK293T cells (ATCC, VA) using polyethylenimine (Polysciences, PEI, 23966-2) and third-generation lentiviral packaging plasmids pMDLg/pRRE (Addgene, 12251), pRSV-RV (Addgene, 12253), and VSC-G (Addgene, 12259), along with pLVX-UbC-rtTA-Ngn2:2A (Addgene, 127288).

    Techniques: Derivative Assay, Transduction, Western Blot, Expressing, Quantitative RT-PCR, Fluorescence, Immunocytochemistry, Cell Culture, Labeling, Control