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plasmid encoding vsvg  (Addgene inc)


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    Addgene inc plasmid encoding vsvg
    <t>HIV</t> infection induces miR-422a in CD4+ T cells via expression of Nef (A) Primary CD4+ T cells were stimulated with anti-CD3/CD28 beads for 3 days, then infected with HIV-1 NL4-3 . The spreading infection was measured by evaluating supernatant p24 concentrations by ELISA at each time point. (B and C) The primary (B) and mature miR-422a expression levels (C) were measured by RT-qPCR after infection with HIV-1 in primary CD4+ T cells, as described in (A). (D) Stimulated primary CD4+ T cells were infected with an HIV-1 single-round reporter virus <t>(VSVG-pseudotyped</t> HIV NL4-3 ΔEnv EGFP reporter virus) for 3 days. Cells were then sorted based on EGFP expression using the Sony MA900 multi-application cell sorter and harvested for RNA isolation. miR-422a expression was measured by RT-qPCR. (E) Stimulated primary CD4+ T cells were infected with HIV-1 single-round reporter virus (VSVG-pseudotyped HIV NL4-3 ΔEnv ΔNef GFP reporter virus) for 3 days. Cells were then sorted based on GFP expression for RNA isolation. miR-422a expression was measured by RT-qPCR. (F) Nef was nucleofected into stimulated primary CD4+ T cells for 48 h. Cells were then collected to evaluate miR-422a expression by RT-qPCR. G. Nef was nucleofected into stimulated primary CD4+ T cells for 48 h. Cells were then stained with anti-CD25 and anti-69 antibodies to detect CD25 and CD69 expression by flow cytometry. Each dot represents data from one donor. Data are representative of the results of three independent experiments ( n = 3 biologically independent samples, mean ± SEM). Statistical significance was analyzed by unpaired or paired student t tests. p ≤ 0.05 [∗], p ≤ 0.01 [∗∗], p ≤ 0.001 [∗∗∗], p ≤ 0.0001 [∗∗∗∗].
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    1) Product Images from "microRNA-422a promotes HIV replication and innate immune evasion by targeting MECP2"

    Article Title: microRNA-422a promotes HIV replication and innate immune evasion by targeting MECP2

    Journal: Molecular Therapy. Nucleic Acids

    doi: 10.1016/j.omtn.2026.102844

    HIV infection induces miR-422a in CD4+ T cells via expression of Nef (A) Primary CD4+ T cells were stimulated with anti-CD3/CD28 beads for 3 days, then infected with HIV-1 NL4-3 . The spreading infection was measured by evaluating supernatant p24 concentrations by ELISA at each time point. (B and C) The primary (B) and mature miR-422a expression levels (C) were measured by RT-qPCR after infection with HIV-1 in primary CD4+ T cells, as described in (A). (D) Stimulated primary CD4+ T cells were infected with an HIV-1 single-round reporter virus (VSVG-pseudotyped HIV NL4-3 ΔEnv EGFP reporter virus) for 3 days. Cells were then sorted based on EGFP expression using the Sony MA900 multi-application cell sorter and harvested for RNA isolation. miR-422a expression was measured by RT-qPCR. (E) Stimulated primary CD4+ T cells were infected with HIV-1 single-round reporter virus (VSVG-pseudotyped HIV NL4-3 ΔEnv ΔNef GFP reporter virus) for 3 days. Cells were then sorted based on GFP expression for RNA isolation. miR-422a expression was measured by RT-qPCR. (F) Nef was nucleofected into stimulated primary CD4+ T cells for 48 h. Cells were then collected to evaluate miR-422a expression by RT-qPCR. G. Nef was nucleofected into stimulated primary CD4+ T cells for 48 h. Cells were then stained with anti-CD25 and anti-69 antibodies to detect CD25 and CD69 expression by flow cytometry. Each dot represents data from one donor. Data are representative of the results of three independent experiments ( n = 3 biologically independent samples, mean ± SEM). Statistical significance was analyzed by unpaired or paired student t tests. p ≤ 0.05 [∗], p ≤ 0.01 [∗∗], p ≤ 0.001 [∗∗∗], p ≤ 0.0001 [∗∗∗∗].
    Figure Legend Snippet: HIV infection induces miR-422a in CD4+ T cells via expression of Nef (A) Primary CD4+ T cells were stimulated with anti-CD3/CD28 beads for 3 days, then infected with HIV-1 NL4-3 . The spreading infection was measured by evaluating supernatant p24 concentrations by ELISA at each time point. (B and C) The primary (B) and mature miR-422a expression levels (C) were measured by RT-qPCR after infection with HIV-1 in primary CD4+ T cells, as described in (A). (D) Stimulated primary CD4+ T cells were infected with an HIV-1 single-round reporter virus (VSVG-pseudotyped HIV NL4-3 ΔEnv EGFP reporter virus) for 3 days. Cells were then sorted based on EGFP expression using the Sony MA900 multi-application cell sorter and harvested for RNA isolation. miR-422a expression was measured by RT-qPCR. (E) Stimulated primary CD4+ T cells were infected with HIV-1 single-round reporter virus (VSVG-pseudotyped HIV NL4-3 ΔEnv ΔNef GFP reporter virus) for 3 days. Cells were then sorted based on GFP expression for RNA isolation. miR-422a expression was measured by RT-qPCR. (F) Nef was nucleofected into stimulated primary CD4+ T cells for 48 h. Cells were then collected to evaluate miR-422a expression by RT-qPCR. G. Nef was nucleofected into stimulated primary CD4+ T cells for 48 h. Cells were then stained with anti-CD25 and anti-69 antibodies to detect CD25 and CD69 expression by flow cytometry. Each dot represents data from one donor. Data are representative of the results of three independent experiments ( n = 3 biologically independent samples, mean ± SEM). Statistical significance was analyzed by unpaired or paired student t tests. p ≤ 0.05 [∗], p ≤ 0.01 [∗∗], p ≤ 0.001 [∗∗∗], p ≤ 0.0001 [∗∗∗∗].

    Techniques Used: Infection, Expressing, Enzyme-linked Immunosorbent Assay, Quantitative RT-PCR, Virus, Isolation, Staining, Flow Cytometry

    Related Articles

    Cotransfection:

    Article Title: Sensing of HIV-1 Entry Triggers a Type I Interferon Response in Human Primary Macrophages
    Article Snippet: HIV-1 NL4-3ΔEnv (kindly provided by O. Schwartz, Institut Pasteur, Paris, France), HIV-1 NL-AD8 ( 50 ), and psPAX2 VLP were produced by transfection of the corresponding proviral DNA or psPAX2 plasmid (12260; Addgene) in HEK 293T cells (ATCC number CRL-11268) by polyethyleneimine (PEI) precipitation. .. Pseudotyping was achieved by cotransfection with a plasmid encoding VSVG (pMD2.G; 12259; Addgene) or ADA Env (kind gift of M. Negroni, IBMC, Strasbourg, France) ( 51 ). ..

    Plasmid Preparation:

    Article Title: Sensing of HIV-1 Entry Triggers a Type I Interferon Response in Human Primary Macrophages
    Article Snippet: HIV-1 NL4-3ΔEnv (kindly provided by O. Schwartz, Institut Pasteur, Paris, France), HIV-1 NL-AD8 ( 50 ), and psPAX2 VLP were produced by transfection of the corresponding proviral DNA or psPAX2 plasmid (12260; Addgene) in HEK 293T cells (ATCC number CRL-11268) by polyethyleneimine (PEI) precipitation. .. Pseudotyping was achieved by cotransfection with a plasmid encoding VSVG (pMD2.G; 12259; Addgene) or ADA Env (kind gift of M. Negroni, IBMC, Strasbourg, France) ( 51 ). ..

    Article Title: Sensing of HIV-1 Entry Triggers a Type I Interferon Response in Human Primary Macrophages
    Article Snippet: 124 HIV-1 NL4-3ΔEnv (kindly provided by O. Schwartz, Institut Pasteur, Paris, France), 125 HIV-1 NL-AD8 (24) and psPAX2 VLP were produced by transfection of the 126 corresponding proviral DNA or psPAX2 plasmid (Addgene #12260) in HEK 293T cells 127 (ATCC number: CRL-11268) by polyethyleneimine (PEI) precipitation. .. Pseudotyping 128 was achieved by co-transfection with a plasmid encoding VSVG (pMD2.G, Addgene 129 #12259), or ADA Env (kind gift of M. Negroni, IBMC, Strasbourg, France) (25). ..

    Article Title: microRNA-422a promotes HIV replication and innate immune evasion by targeting MECP2
    Article Snippet: HIV-1 NL4-3 viral stocks were generated by transfection of proviral DNA (BEI, #ARP-114) into HEK-293T cells via polyethylenimine (PEI, Polysciences, #26913-06-4) as described previously. .. Specifically, VSVG-pseudotyped HIV NL4-3 ΔEnv EGFP reporter virus stocks were produced by co-transfecting the plasmid encoding HIV NL4-3 ΔEnv EGFP (BEI, #ARP-11100) and the plasmid encoding VSVG (Addgene, #8454) at a ratio of 3:1 into HEK-293T cells at 50%–60% confluency in culture flasks. ..

    Virus:

    Article Title: microRNA-422a promotes HIV replication and innate immune evasion by targeting MECP2
    Article Snippet: HIV-1 NL4-3 viral stocks were generated by transfection of proviral DNA (BEI, #ARP-114) into HEK-293T cells via polyethylenimine (PEI, Polysciences, #26913-06-4) as described previously. .. Specifically, VSVG-pseudotyped HIV NL4-3 ΔEnv EGFP reporter virus stocks were produced by co-transfecting the plasmid encoding HIV NL4-3 ΔEnv EGFP (BEI, #ARP-11100) and the plasmid encoding VSVG (Addgene, #8454) at a ratio of 3:1 into HEK-293T cells at 50%–60% confluency in culture flasks. ..

    Produced:

    Article Title: microRNA-422a promotes HIV replication and innate immune evasion by targeting MECP2
    Article Snippet: HIV-1 NL4-3 viral stocks were generated by transfection of proviral DNA (BEI, #ARP-114) into HEK-293T cells via polyethylenimine (PEI, Polysciences, #26913-06-4) as described previously. .. Specifically, VSVG-pseudotyped HIV NL4-3 ΔEnv EGFP reporter virus stocks were produced by co-transfecting the plasmid encoding HIV NL4-3 ΔEnv EGFP (BEI, #ARP-11100) and the plasmid encoding VSVG (Addgene, #8454) at a ratio of 3:1 into HEK-293T cells at 50%–60% confluency in culture flasks. ..



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    Thermo Fisher plp1, plp2, plp/vsvg plasmids encoding viral proteins gag-pol, rev, vsv-g
    <t>HIV</t> infection induces miR-422a in CD4+ T cells via expression of Nef (A) Primary CD4+ T cells were stimulated with anti-CD3/CD28 beads for 3 days, then infected with HIV-1 NL4-3 . The spreading infection was measured by evaluating supernatant p24 concentrations by ELISA at each time point. (B and C) The primary (B) and mature miR-422a expression levels (C) were measured by RT-qPCR after infection with HIV-1 in primary CD4+ T cells, as described in (A). (D) Stimulated primary CD4+ T cells were infected with an HIV-1 single-round reporter virus <t>(VSVG-pseudotyped</t> HIV NL4-3 ΔEnv EGFP reporter virus) for 3 days. Cells were then sorted based on EGFP expression using the Sony MA900 multi-application cell sorter and harvested for RNA isolation. miR-422a expression was measured by RT-qPCR. (E) Stimulated primary CD4+ T cells were infected with HIV-1 single-round reporter virus (VSVG-pseudotyped HIV NL4-3 ΔEnv ΔNef GFP reporter virus) for 3 days. Cells were then sorted based on GFP expression for RNA isolation. miR-422a expression was measured by RT-qPCR. (F) Nef was nucleofected into stimulated primary CD4+ T cells for 48 h. Cells were then collected to evaluate miR-422a expression by RT-qPCR. G. Nef was nucleofected into stimulated primary CD4+ T cells for 48 h. Cells were then stained with anti-CD25 and anti-69 antibodies to detect CD25 and CD69 expression by flow cytometry. Each dot represents data from one donor. Data are representative of the results of three independent experiments ( n = 3 biologically independent samples, mean ± SEM). Statistical significance was analyzed by unpaired or paired student t tests. p ≤ 0.05 [∗], p ≤ 0.01 [∗∗], p ≤ 0.001 [∗∗∗], p ≤ 0.0001 [∗∗∗∗].
    Plp1, Plp2, Plp/Vsvg Plasmids Encoding Viral Proteins Gag Pol, Rev, Vsv G, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 90/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    HIV infection induces miR-422a in CD4+ T cells via expression of Nef (A) Primary CD4+ T cells were stimulated with anti-CD3/CD28 beads for 3 days, then infected with HIV-1 NL4-3 . The spreading infection was measured by evaluating supernatant p24 concentrations by ELISA at each time point. (B and C) The primary (B) and mature miR-422a expression levels (C) were measured by RT-qPCR after infection with HIV-1 in primary CD4+ T cells, as described in (A). (D) Stimulated primary CD4+ T cells were infected with an HIV-1 single-round reporter virus (VSVG-pseudotyped HIV NL4-3 ΔEnv EGFP reporter virus) for 3 days. Cells were then sorted based on EGFP expression using the Sony MA900 multi-application cell sorter and harvested for RNA isolation. miR-422a expression was measured by RT-qPCR. (E) Stimulated primary CD4+ T cells were infected with HIV-1 single-round reporter virus (VSVG-pseudotyped HIV NL4-3 ΔEnv ΔNef GFP reporter virus) for 3 days. Cells were then sorted based on GFP expression for RNA isolation. miR-422a expression was measured by RT-qPCR. (F) Nef was nucleofected into stimulated primary CD4+ T cells for 48 h. Cells were then collected to evaluate miR-422a expression by RT-qPCR. G. Nef was nucleofected into stimulated primary CD4+ T cells for 48 h. Cells were then stained with anti-CD25 and anti-69 antibodies to detect CD25 and CD69 expression by flow cytometry. Each dot represents data from one donor. Data are representative of the results of three independent experiments ( n = 3 biologically independent samples, mean ± SEM). Statistical significance was analyzed by unpaired or paired student t tests. p ≤ 0.05 [∗], p ≤ 0.01 [∗∗], p ≤ 0.001 [∗∗∗], p ≤ 0.0001 [∗∗∗∗].

    Journal: Molecular Therapy. Nucleic Acids

    Article Title: microRNA-422a promotes HIV replication and innate immune evasion by targeting MECP2

    doi: 10.1016/j.omtn.2026.102844

    Figure Lengend Snippet: HIV infection induces miR-422a in CD4+ T cells via expression of Nef (A) Primary CD4+ T cells were stimulated with anti-CD3/CD28 beads for 3 days, then infected with HIV-1 NL4-3 . The spreading infection was measured by evaluating supernatant p24 concentrations by ELISA at each time point. (B and C) The primary (B) and mature miR-422a expression levels (C) were measured by RT-qPCR after infection with HIV-1 in primary CD4+ T cells, as described in (A). (D) Stimulated primary CD4+ T cells were infected with an HIV-1 single-round reporter virus (VSVG-pseudotyped HIV NL4-3 ΔEnv EGFP reporter virus) for 3 days. Cells were then sorted based on EGFP expression using the Sony MA900 multi-application cell sorter and harvested for RNA isolation. miR-422a expression was measured by RT-qPCR. (E) Stimulated primary CD4+ T cells were infected with HIV-1 single-round reporter virus (VSVG-pseudotyped HIV NL4-3 ΔEnv ΔNef GFP reporter virus) for 3 days. Cells were then sorted based on GFP expression for RNA isolation. miR-422a expression was measured by RT-qPCR. (F) Nef was nucleofected into stimulated primary CD4+ T cells for 48 h. Cells were then collected to evaluate miR-422a expression by RT-qPCR. G. Nef was nucleofected into stimulated primary CD4+ T cells for 48 h. Cells were then stained with anti-CD25 and anti-69 antibodies to detect CD25 and CD69 expression by flow cytometry. Each dot represents data from one donor. Data are representative of the results of three independent experiments ( n = 3 biologically independent samples, mean ± SEM). Statistical significance was analyzed by unpaired or paired student t tests. p ≤ 0.05 [∗], p ≤ 0.01 [∗∗], p ≤ 0.001 [∗∗∗], p ≤ 0.0001 [∗∗∗∗].

    Article Snippet: Specifically, VSVG-pseudotyped HIV NL4-3 ΔEnv EGFP reporter virus stocks were produced by co-transfecting the plasmid encoding HIV NL4-3 ΔEnv EGFP (BEI, #ARP-11100) and the plasmid encoding VSVG (Addgene, #8454) at a ratio of 3:1 into HEK-293T cells at 50%–60% confluency in culture flasks.

    Techniques: Infection, Expressing, Enzyme-linked Immunosorbent Assay, Quantitative RT-PCR, Virus, Isolation, Staining, Flow Cytometry