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526 results sorted by
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As the stability and half-life of various mRNAs and their protein products varies, it is important to empirically determine the best time points for assessing target knockdown. For example, it has been documented that in mammalian cells, mRNA...
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The media recommended for these cells is Leibovitz's L-15 medium which does not contain sodium bicarbonate to act as a buffer for the carbonic acid that would normally form in the presence of CO2. Furthermore, if the cells are cultured in Leibovitz's...
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Depending on individual cell type and doubling rate, selection of stable transfectants will take between 7 and 28 days. Expansion and characterization of single cell clones will take several weeks in addition.
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This is the normal pattern you should expect to see. Only a small proportion (1/10,000 to 1/100) of all transfected cells will integrate the transfected DNA into their genome and become stable transfectants. The remaining cells lose the transfected...
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Yes. Please click on the attached Technical Reference Guide: "Designing an RNAi Experiment Using Nucleofection®". On page 3 you can find a table with some examples.
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Yes. As the same protocol applies for any nucleic acid substrate (vectors or oligonucleotides) you can easily perform co-transfections either for transfection control or rescue experiments.
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The most commonly used marker is the neomycin phosphotransferase (neo) gene that confers resistance to G418 to eukaryotic cells. Other markers are Puromycin, Hygromycin, Zeocin, or the HPRT gene that can be used in HPRT-deficient cells.
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Yes, the optimized protocols for the standard Nucleofector® recommend the use of 6 well plates. Plating into a 12-well plate will pose a problem as less cells will attach at high densities.
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Antibiotics for selection should be added 24-48 h after Nucleofection®.
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We recommend using cuvettes with the 0.1 cm gap width.
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