The proven activated dendrimer for transfection of a broad range of cell lines
Serum and DNA Quantity vs. Transfection Efficiency
Influence of serum and DNA quantity on transfection using SuperFect Reagent. 2 x 104 COS-7 cells were seeded per well in 96-well plates one day prior transfection. Cells were transfected using 0.1每2.0 µg of a beta-galactosidase每reporter plasmid and 3 µl SuperFect Reagent per well, in either the presence or absence of serum. Each bar represents the average efficiency from four replicates assayed 48 h post-transfection.
SuperFect Reagent consists of activated-dendrimer molecules with a defined spherical architecture (1). Branches radiate from a central core and terminate at charged amino groups which can then interact with negatively charged phosphate groups of nucleic acids (see figure "Activated-Dendrimer Structure"). SuperFect Reagent assembles DNA into compact structures (see figure "SuperFect每DNA Interaction") that bind to the cell surface and are taken into the cell by nonspecific endocytosis. The reagent buffers the pH of the endosome, leading to pH inhibition of endosomal nucleases, which ensures stability of SuperFect每DNA complexes.
Due to highly controlled chemical synthesis the activated-dendrimer molecules in SuperFect Reagent have a precise size and a defined shape. This ensures consistent transfection-complex formation and reproducible transfection results.
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Schematic representation of an activated dendrimer (1). Note the highly branched structure. |
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Model of the SuperFect每DNA complex. SuperFect Reagent (purple balls) interacts with DNA (black) to form a ring-like (toroid-like) structure. The upper right section of the illustration shows naked DNA, the lower section shows the interaction between dendrimers and DNA inside the complex, and the upper left section shows the final complete coverage of DNA within the complex. |
SuperFect Transfection Reagent is provided as a ready-to-use solution. The reagent is simply added to the DNA solution, mixed, and incubated for 5每10 minutes to allow SuperFect每DNA complexes to form. The complexes are mixed with growth medium (which can contain serum and antibiotics), and added directly to the cells. After a 2每3 hour incubation, a medium change is performed and the cells are incubated for expression of the transfected gene.
SuperFect Reagent is suitable for transient and stable transfection of a broad range of cell lines (see figure "Transfection of Neuronal PC-12 Cells Using SuperFect Reagent"). A searchable list of cell lines successfully transfected using SuperFect Reagent, as well as customer-developed transfection protocols, is available at the Transfection Tools web site 〞 www.qiagen.com/transfectiontools/.
Transfection of Neuronal PC-12 Cells Using SuperFect Reagent
Expression of green fluorescent protein (GFP) in differentiated PC-12 cells 5 days post-transfection. 104每105 cells previously stimulated with 50 ng/ml NGF were plated per 60 mm dish one day prior to transfection. Transient transfections were performed in 2 ml low-serum growth medium (DMEM plus 0.05% FBS) using 3 µg of a GFP-reporter plasmid and 15 µl SuperFect Reagent. (Data kindly provided by K. Kelly-Spratt, University of Texas Southwestern Medical Center, Dallas, TX, USA.)
The application of recombinant DNA technology to fields such as drug discovery and development has led to an increased need for high-throughput transfection. Transfection using SuperFect Reagent requires minimal handling, making it highly suitable for high-throughput screening. SuperFect Reagent provides high transfection efficiencies, excellent reproducibility, and low cytotoxicity in high-throughput transfection, and is available in bulk quantities.
紱釬忒聊:Superfect Transfection Reagent Handbook 12/00 (PDF version, 121 KB)
1. Tang, M.X., Redemann, C.T., and Szoka, Jr., F.C. (1996) In vitro gene delivery by degraded polyamidoamine dendrimers. Bioconjugate Chem. 7, 703.
For a complete list of references citing uses of QIAGEN products, visit the QIAGEN Reference Database or contact QIAGEN Technical Services or your local distributor.