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TransMessenger Transfection Reagent Handbook

NA preparations can potentially lower transfection efficiency. Degradation of RNA leads to decreased expression levels. RNA should be checked for degradation before transfection. We recommend purifying RNA using RNeasy Kits, Oligotex mRNA Kits, or QIAGEN DNA/RNA Kits. RNA-related effects Toxic effects may arise if RNA encoding a toxic protein is transfected, or if too much RNA is used. In contrast, if insufficient RNA with a low translation rate is used, transfection efficiency may be too low. Optimize the amount of RNA according to the optimization guide for each new RNA and/or cell type used. Key gene is silenced If the gene targeted in an RNAi experiment is key to the survival of the cell, silencing this gene will lead to cell death. Variable
transfection
efficiencies in
replicate
experiments
Inconsistent cell
confluencies in
replicate experiments Count cells prior to seeding to ensure that the same number of cells is seeded for each
experiment. Keep incubation times between seeding and complex addition consistent
between experiments. Cells should be seeded no later than 24 h before transfection. Possible mycoplasma
contamination Mycoplasma contamination influences transfection efficiency. Variations in the growth behavior of mycoplasma-infected cells will lead to different transfection efficiencies between replicate experiments.
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