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Effects of Circular DNA Length on Transfection Efficiency by Electroporation into HeLa Cells
The ability to produce extremely small and circular supercoiled vectors has opened new territory for improving non-viral gene therapy vectors. In this work, we compared transfection of supercoiled DNA vectors ranging from 383 to 4,548 bp, each encoding shRNA against GFP under control of the H1 promo...
Autores principales: | , , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
Public Library of Science
2016
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5137892/ https://www.ncbi.nlm.nih.gov/pubmed/27918590 http://dx.doi.org/10.1371/journal.pone.0167537 |
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author | Hornstein, Benjamin D. Roman, Dany Arévalo-Soliz, Lirio M. Engevik, Melinda A. Zechiedrich, Lynn |
author_facet | Hornstein, Benjamin D. Roman, Dany Arévalo-Soliz, Lirio M. Engevik, Melinda A. Zechiedrich, Lynn |
author_sort | Hornstein, Benjamin D. |
collection | PubMed |
description | The ability to produce extremely small and circular supercoiled vectors has opened new territory for improving non-viral gene therapy vectors. In this work, we compared transfection of supercoiled DNA vectors ranging from 383 to 4,548 bp, each encoding shRNA against GFP under control of the H1 promoter. We assessed knockdown of GFP by electroporation into HeLa cells. All of our vectors entered cells in comparable numbers when electroporated with equal moles of DNA. Despite similar cell entry, we found length-dependent differences in how efficiently the vectors knocked down GFP. As vector length increased up to 1,869 bp, GFP knockdown efficiency per mole of transfected DNA increased. From 1,869 to 4,257 bp, GFP knockdown efficiency per mole was steady, then decreased with increasing vector length. In comparing GFP knockdown with equal masses of vectors, we found that the shorter vectors transfect more efficiently per nanogram of DNA transfected. Our results rule out cell entry and DNA mass as determining factors for gene knockdown efficiency via electroporation. The length-dependent effects we have uncovered are likely explained by differences in nuclear translocation or transcription. These data add an important step towards clinical applications of non-viral vector delivery. |
format | Online Article Text |
id | pubmed-5137892 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2016 |
publisher | Public Library of Science |
record_format | MEDLINE/PubMed |
spelling | pubmed-51378922016-12-21 Effects of Circular DNA Length on Transfection Efficiency by Electroporation into HeLa Cells Hornstein, Benjamin D. Roman, Dany Arévalo-Soliz, Lirio M. Engevik, Melinda A. Zechiedrich, Lynn PLoS One Research Article The ability to produce extremely small and circular supercoiled vectors has opened new territory for improving non-viral gene therapy vectors. In this work, we compared transfection of supercoiled DNA vectors ranging from 383 to 4,548 bp, each encoding shRNA against GFP under control of the H1 promoter. We assessed knockdown of GFP by electroporation into HeLa cells. All of our vectors entered cells in comparable numbers when electroporated with equal moles of DNA. Despite similar cell entry, we found length-dependent differences in how efficiently the vectors knocked down GFP. As vector length increased up to 1,869 bp, GFP knockdown efficiency per mole of transfected DNA increased. From 1,869 to 4,257 bp, GFP knockdown efficiency per mole was steady, then decreased with increasing vector length. In comparing GFP knockdown with equal masses of vectors, we found that the shorter vectors transfect more efficiently per nanogram of DNA transfected. Our results rule out cell entry and DNA mass as determining factors for gene knockdown efficiency via electroporation. The length-dependent effects we have uncovered are likely explained by differences in nuclear translocation or transcription. These data add an important step towards clinical applications of non-viral vector delivery. Public Library of Science 2016-12-05 /pmc/articles/PMC5137892/ /pubmed/27918590 http://dx.doi.org/10.1371/journal.pone.0167537 Text en © 2016 Hornstein et al http://creativecommons.org/licenses/by/4.0/ This is an open access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/4.0/) , which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. |
spellingShingle | Research Article Hornstein, Benjamin D. Roman, Dany Arévalo-Soliz, Lirio M. Engevik, Melinda A. Zechiedrich, Lynn Effects of Circular DNA Length on Transfection Efficiency by Electroporation into HeLa Cells |
title | Effects of Circular DNA Length on Transfection Efficiency by Electroporation into HeLa Cells |
title_full | Effects of Circular DNA Length on Transfection Efficiency by Electroporation into HeLa Cells |
title_fullStr | Effects of Circular DNA Length on Transfection Efficiency by Electroporation into HeLa Cells |
title_full_unstemmed | Effects of Circular DNA Length on Transfection Efficiency by Electroporation into HeLa Cells |
title_short | Effects of Circular DNA Length on Transfection Efficiency by Electroporation into HeLa Cells |
title_sort | effects of circular dna length on transfection efficiency by electroporation into hela cells |
topic | Research Article |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5137892/ https://www.ncbi.nlm.nih.gov/pubmed/27918590 http://dx.doi.org/10.1371/journal.pone.0167537 |
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