ON-TARGETplus Non-targeting Pool

Negative control pool of four siRNAs designed and microarray tested for minimal targeting of human, mouse or rat genes. ON-TARGETplus modifications reduce potential off-targets. Recommended for determination of baseline cellular responses in RNAi experiments.

ON-TARGETplus Non-targeting Control Pool is the ideal negative control for RNAi experiments in human, mouse, and rat cells. The four component ON-TARGETplus Non-targeting siRNAs were designed to leverage seed-region optimization and patented ON-TARGETplus modification patterns to have fewer off-targets than traditionally designed negative control siRNAs, and screened by genomewide microarray analysis. Changes in mRNA or protein levels in cells treated with these controls reflect a baseline cellular response that can be compared to the levels in cells treated with target-specific siRNA.

Highlights

• Ideal negative control for RNAi experiments in human, mouse, and rat cells
• Chemically matched for use with ON-TARGETplus siRNA reagents
• Screened by genome-wide microarray analysis for minimal off-targets
• Double-stranded, chemically synthesized oligonucleotides
• Both sense and antisense strands contain UU for 3'-overhangs
• Mass of each strand confirmed by MALDI-TOF mass spectrometry
• Duplex integrity confirmed by non-denaturing polyacrylamide gel electrophoresis

RNA samples from HeLa cells were analyzed by microarray expression profiling 24 hours after transfection with the indicated siRNA reagents. Decreases in mRNA levels are shown in green and increases in red. The left panel shows a typical off-target signature of a “Scramble” control siRNA. The right panel shows the off-targets for ON-TARGETplus Non-Targeting Control siRNAs 1 through 4. (D-001810-01, -02, 03, 04).

Storage
siRNA should be resuspended in RNase-free solutions. We recommend 1x siRNA buffer (diluted from 5x siRNA buffer – Dharmacon
Cat. #B-002000-UB-100). RNase-free water (for short-term storage) is also appropriate for resuspension of concentrated stocks (20-100 μM).
Alternatively, an RNase-free buffer (pH 7.3-7.6) may be used such as PBS (Dharmacon Cat. #NC9826748).  

Upon resuspension, aliquot the siRNA into small volumes and store at -20 °C to -80 °C. For best results, limit freeze-thawing of each tube to no more than five events. Under these conditions, the siRNA is stable for at least 6 months.

Citations
P. Baum, K. Fundel-Clemens, S. Kreuz, R. E. Kontermann, A. Weith, D. Mennerich, J. F. Rippmann. Off-Target Analysis of Control siRNA Molecules Reveals Important Differences in the Cytokine Profile and Inflammation Response of Human Fibroblasts. Oligonucleotides. 20(1), 17-25 (2010).