Document Detail


The effects of plasmid copy number and sequence context upon transfection efficiency.
MedLine Citation:
PMID:  14684288     Owner:  NLM     Status:  MEDLINE    
Abstract/OtherAbstract:
It is known that large P1 artificial chromosome (PAC) vectors exhibit reduced transfection efficiency in comparison to small plasmid vectors. We investigated the dynamics of this effect, by comparing expression from a small plasmid (4.7 kb) and a PAC vector (111 kb) containing the Enhanced Green Fluorescent Protein (EGFP) reporter gene under the control of a P(CMV) promoter. EGFP expression was detected by fluorescence activated cell sorting (FACS). We found that the lower transfection efficiency of PAC vectors represents both a smaller percentage of cells expressing the transgene, and a lower level of expression per cell. We have shown that the lower number of plasmid molecules administered per cell in a PAC transfection does not explain this effect, and that this effect does not act in trans. Surprisingly, dilution of a reporter construct with an irrelevant plasmid did not appear to compromise transfection efficiency; in fact, a dilution of 1/10 slightly enhanced transfection. Therefore, it seems that the plasmid content of a liposome-DNA complex need not be 100% reporter construct for optimum transfection efficiency. This discovery has potential practical utility in a number of applications.
Authors:
Wendy E Walker; David J Porteous; A Christopher Boyd
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Publication Detail:
Type:  Journal Article; Research Support, Non-U.S. Gov't    
Journal Detail:
Title:  Journal of controlled release : official journal of the Controlled Release Society     Volume:  94     ISSN:  0168-3659     ISO Abbreviation:  J Control Release     Publication Date:  2004 Jan 
Date Detail:
Created Date:  2003-12-19     Completed Date:  2004-09-08     Revised Date:  2006-11-15    
Medline Journal Info:
Nlm Unique ID:  8607908     Medline TA:  J Control Release     Country:  Netherlands    
Other Details:
Languages:  eng     Pagination:  245-52     Citation Subset:  IM    
Affiliation:
Molecular Medicine Centre, Department of Medicine, University of Edinburgh, Western General Hospital, Crewe Road, EH4 2XU, UK.
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MeSH Terms
Descriptor/Qualifier:
Animals
Base Sequence
COS Cells
Cercopithecus aethiops
Gene Dosage*
Genetic Vectors
Plasmids / administration & dosage,  genetics,  pharmacokinetics*
Transfection / methods*

From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine


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