Lipofectamine® 2000 Transfection Reagent

Protocol tips

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	Add diluted siRNA to diluted Lipofectamine Reagent Incubate for 5 min at RT and add to cells. Incubate cells for 1–3 days at 37°C.

Protocol tips
Add diluted siRNA to diluted Lipofectamine Reagent Incubate for 5 min at RT and add to cells. Incubate cells for 1–3 days at 37°C.

Publication protocol

Calcium phosphate nanoparticles were precipitated from aqueous solutions of calcium nitrate (6.25 mm) and diammonium hydrogen phosphate (3.74 mm) and then functionalized with DNA or siRNA (1 g l−1) and PEI (25 kDa, 2 g l−1) as described earlier75 with the following modifications: CaP/DNA/CaP/PEI nanoparticles were separated from nonadsorbed molecules (including excess nucleic acid or PEI) by centrifugation (5000 g, 4 °C, 15 min) and then dispersed in the half of the original volume of water by ultrasonication (UP50H, Hielscher, Teltow, Germany; Ultrasound Technology; sonotrode 2, cycle 0.8, amplitude 80%, 10 s). CaP/siRNA/CaP/PEI nanoparticles were isolated by centrifugation (20 000 g, 4 °C, 15 min) and then dispersed in the original volume of water by ultrasonication with the same conditions.

Model DNA leading to the expression of eGFP (pcDNA3-eGFP) was used. For the gene-silencing experiments the eGFP-siRNA: sense, 5′-GCAAGCUGACCCUGAAGUUCAU-3′ antisense, 5′-AUGAACUUCAGGGUCAGCUUGC-3′ (Invitrogen, Paisley, UK) was used. Lipofectamine 2000 was obtained from Life Technologies (Invitrogen, Carlsbad, CA, USA) and used according to the manufacturer's specifications.

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Manufacturer protocol

Download the product protocol from Thermo Fisher Scientific for Lipofectamine® 2000 Transfection Reagent below.

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