Abstract
A bi-functional, 31 amino acid synthetic peptide (polylysine-molossin) was evaluated for gene delivery to primary cultures of rat cerebral cortex neurons. Polylysine-molossin consists of an amino terminal domain of 16 lysines for electrostatic binding of DNA, and a 15 amino acid, integrin-binding domain at the carboxyl terminal. High levels of gene delivery were obtained with 20-30 microM chloroquine, with a synthetic fusogenic peptide at an optimal DNA:polylysine-molossin:fusogenic peptide w/w ratio of 1:3:0.2, and with the addition of low concentrations of Lipofectamine 2000 at an optimal DNA:polylysine-molossin:Lipofectamine 2000 w/w ratio of 1:3:0.5. With the best combination, >30% of neurons strongly expressed the beta-galactosidase reporter gene, with no observable toxicity. DNA concentrations >2 microgram/ml were essential for efficient gene delivery. This synthetic peptide provides a safe, readily standardised and flexible DNA vector system well suited to ex vivo gene delivery to neurons for experimental and clinical applications.
Publication types
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Comparative Study
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Research Support, Non-U.S. Gov't
MeSH terms
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Animals
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Antirheumatic Agents / pharmacology
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Carrier Proteins / genetics*
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Cation Exchange Resins / pharmacology
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Cells, Cultured
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Cerebral Cortex / cytology
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Chloroquine / pharmacology
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Crotalid Venoms / genetics*
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Dose-Response Relationship, Drug
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Drug Interactions
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Female
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Fetus
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Gene Expression
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Gene Transfer Techniques
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Genes, Reporter / drug effects
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Genes, Reporter / physiology
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Genetic Vectors / genetics
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Glycoside Hydrolases*
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Hemagglutinins
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Hemagglutinins, Viral*
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Indicators and Reagents / pharmacology
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Lipids / pharmacology
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Luciferases / metabolism
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Neurons / physiology*
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Oligopeptides / genetics*
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Oligopeptides / metabolism
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Peptide Fragments / genetics*
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Plasmids / genetics
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Pregnancy
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Rats
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Rats, Sprague-Dawley
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Transfection / methods*
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Viral Proteins
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beta-Galactosidase / biosynthesis
Substances
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Antirheumatic Agents
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Carrier Proteins
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Cation Exchange Resins
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Crotalid Venoms
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Hemagglutinins
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Hemagglutinins, Viral
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Indicators and Reagents
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Lipids
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Lipofectamine
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Oligopeptides
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Peptide Fragments
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Viral Proteins
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hemagglutinin fusogenic peptide, influenza virus
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polylysine-molossin peptide
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Chloroquine
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Luciferases
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Glycoside Hydrolases
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beta-galactanase
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beta-Galactosidase