Efficient gene delivery to primary neuron cultures using a synthetic peptide vector system

Collins, Louise, Asuni, Ayodeji A., Anderton, Brian H. and Fabre, John W. (2003) Efficient gene delivery to primary neuron cultures using a synthetic peptide vector system. Journal of Neuroscience Methods, 125 (1-2), 113-120. (doi:10.1016/S0165-0270(03)00042-6).

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 ?M 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 ?-galactosidase reporter gene, with no observable toxicity. DNA concentrations >2 ?g/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.

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