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Efficient Transfection of Large Plasmids Encoding HIV-1 into Human Cells—A High Potential Transfection System Based on a Peptide Mimicking Cationic Lipid
One major disadvantage of nucleic acid delivery systems is the low transfection or transduction efficiency of large-sized plasmids into cells. In this communication, we demonstrate the efficient transfection of a 15.5 kb green fluorescent protein (GFP)-fused HIV-1 molecular clone with a nucleic acid...
Autores principales: | , , , , , , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
MDPI
2020
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7559901/ https://www.ncbi.nlm.nih.gov/pubmed/32854383 http://dx.doi.org/10.3390/pharmaceutics12090805 |
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author | Janich, Christopher Ivanusic, Daniel Giselbrecht, Julia Janich, Elena Pinnapireddy, Shashank Reddy Hause, Gerd Bakowsky, Udo Langner, Andreas Wölk, Christian |
author_facet | Janich, Christopher Ivanusic, Daniel Giselbrecht, Julia Janich, Elena Pinnapireddy, Shashank Reddy Hause, Gerd Bakowsky, Udo Langner, Andreas Wölk, Christian |
author_sort | Janich, Christopher |
collection | PubMed |
description | One major disadvantage of nucleic acid delivery systems is the low transfection or transduction efficiency of large-sized plasmids into cells. In this communication, we demonstrate the efficient transfection of a 15.5 kb green fluorescent protein (GFP)-fused HIV-1 molecular clone with a nucleic acid delivery system prepared from the highly potent peptide-mimicking cationic lipid OH4 in a mixture with the phospholipid DOPE (co-lipid). For the transfection, liposomes were loaded using a large-sized plasmid (15.5 kb), which encodes a replication-competent HIV type 1 molecular clone that carries a Gag-internal green fluorescent protein (HIV-1 JR-FL Gag-iGFP). The particle size and charge of the generated nanocarriers with 15.5 kb were compared to those of a standardized 4.7 kb plasmid formulation. Stable, small-sized lipoplexes could be generated independently of the length of the used DNA. The transfer of fluorescently labeled pDNA-HIV1-Gag-iGFP in HEK293T cells was monitored using confocal laser scanning microscopy (cLSM). After efficient plasmid delivery, virus particles were detectable as budding structures on the plasma membrane. Moreover, we observed a randomized distribution of fluorescently labeled lipids over the plasma membrane. Obviously, a significant exchange of lipids between the drug delivery system and the cellular membranes occurs, which hints toward a fusion process. The mechanism of membrane fusion for the internalization of lipid-based drug delivery systems into cells is still a frequently discussed topic. |
format | Online Article Text |
id | pubmed-7559901 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2020 |
publisher | MDPI |
record_format | MEDLINE/PubMed |
spelling | pubmed-75599012020-10-22 Efficient Transfection of Large Plasmids Encoding HIV-1 into Human Cells—A High Potential Transfection System Based on a Peptide Mimicking Cationic Lipid Janich, Christopher Ivanusic, Daniel Giselbrecht, Julia Janich, Elena Pinnapireddy, Shashank Reddy Hause, Gerd Bakowsky, Udo Langner, Andreas Wölk, Christian Pharmaceutics Communication One major disadvantage of nucleic acid delivery systems is the low transfection or transduction efficiency of large-sized plasmids into cells. In this communication, we demonstrate the efficient transfection of a 15.5 kb green fluorescent protein (GFP)-fused HIV-1 molecular clone with a nucleic acid delivery system prepared from the highly potent peptide-mimicking cationic lipid OH4 in a mixture with the phospholipid DOPE (co-lipid). For the transfection, liposomes were loaded using a large-sized plasmid (15.5 kb), which encodes a replication-competent HIV type 1 molecular clone that carries a Gag-internal green fluorescent protein (HIV-1 JR-FL Gag-iGFP). The particle size and charge of the generated nanocarriers with 15.5 kb were compared to those of a standardized 4.7 kb plasmid formulation. Stable, small-sized lipoplexes could be generated independently of the length of the used DNA. The transfer of fluorescently labeled pDNA-HIV1-Gag-iGFP in HEK293T cells was monitored using confocal laser scanning microscopy (cLSM). After efficient plasmid delivery, virus particles were detectable as budding structures on the plasma membrane. Moreover, we observed a randomized distribution of fluorescently labeled lipids over the plasma membrane. Obviously, a significant exchange of lipids between the drug delivery system and the cellular membranes occurs, which hints toward a fusion process. The mechanism of membrane fusion for the internalization of lipid-based drug delivery systems into cells is still a frequently discussed topic. MDPI 2020-08-25 /pmc/articles/PMC7559901/ /pubmed/32854383 http://dx.doi.org/10.3390/pharmaceutics12090805 Text en © 2020 by the authors. Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (http://creativecommons.org/licenses/by/4.0/). |
spellingShingle | Communication Janich, Christopher Ivanusic, Daniel Giselbrecht, Julia Janich, Elena Pinnapireddy, Shashank Reddy Hause, Gerd Bakowsky, Udo Langner, Andreas Wölk, Christian Efficient Transfection of Large Plasmids Encoding HIV-1 into Human Cells—A High Potential Transfection System Based on a Peptide Mimicking Cationic Lipid |
title | Efficient Transfection of Large Plasmids Encoding HIV-1 into Human Cells—A High Potential Transfection System Based on a Peptide Mimicking Cationic Lipid |
title_full | Efficient Transfection of Large Plasmids Encoding HIV-1 into Human Cells—A High Potential Transfection System Based on a Peptide Mimicking Cationic Lipid |
title_fullStr | Efficient Transfection of Large Plasmids Encoding HIV-1 into Human Cells—A High Potential Transfection System Based on a Peptide Mimicking Cationic Lipid |
title_full_unstemmed | Efficient Transfection of Large Plasmids Encoding HIV-1 into Human Cells—A High Potential Transfection System Based on a Peptide Mimicking Cationic Lipid |
title_short | Efficient Transfection of Large Plasmids Encoding HIV-1 into Human Cells—A High Potential Transfection System Based on a Peptide Mimicking Cationic Lipid |
title_sort | efficient transfection of large plasmids encoding hiv-1 into human cells—a high potential transfection system based on a peptide mimicking cationic lipid |
topic | Communication |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7559901/ https://www.ncbi.nlm.nih.gov/pubmed/32854383 http://dx.doi.org/10.3390/pharmaceutics12090805 |
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