Using the right controls in each and every RNAi experiment can speed up your research, streamline analysis, and give you confidence in your results. Invitrogen has a wide selection of controls for your RNAi experiments.
Highly efficient delivery of BLOCK-iT™ Pol II miR RNAi Expression Vectors or BLOCK-iT™ shRNA Vectors is necessary to achieve significant levels of knockdown. Optimizing transfection or transduction, controlling for experimental variability, and using a powerful transfection reagent vastly improves the chances for RNAi success. The use of positive and negative controls may also help in the assessment of your experiments.
Optimizing Transfection/Transduction Conditions
Good results in any RNAi experiment depend upon high transfection or transduction efficiency. Before starting RNAi experiments in a new cell line we recommend first optimizing the transfection/transduction conditions. Invitrogen has a number of free transfection protocols on our website for you to try, but each assay is different and you will probably need to adjust the protocol for your individual needs.
For gene knockdown to be measurable in a cell population, it is very important to have the highest efficiency as possible. Even small reductions in transfection/transduction efficiency can determine whether you can identify functional differences in your experimental samples.
Optimizing delivery conditions
- A DNA plasmid that expresses a fluorescent protein such as GFP monitor that the DNA plasmid has entered the cell
- Dead Cell Stain to monitor toxicity of the transfection
- Nuclei stain to measure the percent of transfected and/or dead cells to the total cell population
- Positive and negative RNAi Vector controls to measure the level of knockdown using each transfection condition.
Monitor experiment-to-experiment transfection/transduction variation.