What is the use of doing siRNA transfection with NC scrambled siRNA for negative control cells? Why we just don't transfect them since they are just a negative control?
Hello Sarah. In general, exogenous DNA or RNA have an impact resulting in a cell response. To ensure that the effect of a specific siRNA transfection is due to its specificity, a control is needed where you use transfection reagents AND a siRNA whose sequence is different from your siRNA of interest. In addition, because in a negative control one doesn't want to observe any effect due to silencing of a specific target, one uses a scrambled siRNA, which is not expected to specifically target any mRNA beyond a background level. Hope this answers your question and clarifies your doubt.
you need a negative control siRNA because it will also be loaded into the cellular RISC. This may compete with endogenous RISC substrates such as miRNAs. A transfection reagent only control will not adequately control for the effects of siRNA loading on RISC.
See https://www.ncbi.nlm.nih.gov/pmc/articles/PMC2782465/ for example.
Hello Sarah. In general, exogenous DNA or RNA have an impact resulting in a cell response. To ensure that the effect of a specific siRNA transfection is due to its specificity, a control is needed where you use transfection reagents AND a siRNA whose sequence is different from your siRNA of interest. In addition, because in a negative control one doesn't want to observe any effect due to silencing of a specific target, one uses a scrambled siRNA, which is not expected to specifically target any mRNA beyond a background level. Hope this answers your question and clarifies your doubt.