Minimizing Off-Target Effects with a Stringent Design Algorithm
Our stringent computer algorithm minimizes off-target effects by using the Smith-Waterman sequence alignment algorithm in a sophisticated filter to design gene-specific siRNA. This algorithm is better than the traditional BLAST search when working with shorter sequences, like siRNA. It insures that a siRNA designed for a specific gene has a sufficient number of mismatches with the rest of the genome to lack an off-target effect. Our siRNA design algorithm also includes filters for all of the published physical and sequence properties known to be important for siRNA activity.

Validated & Guaranteed Knock Down for Each Gene on the Array
Each siRNA pair is validated experimentally on the bench by a well-controlled, rigorous real-time RT-PCR assay insuring greater than 70 percent knock down at the RNA level and thereby minimizing false negatives. You can be sure that the lack of a phenotype for a given gene is NOT due to a lack of gene knockdown. Read more about our validation process in our Validating RNAi White Paper.

Figure: The Knockdown by Each Gene-Specific siRNA Pair in Every Array is Experimentally Validated.
A pair of gene-specific siRNA sequences were designed by the SureSilencing algorithm for 84 different genes. Triplicate reverse transfections of A549, HEK293H, and/or MDA-MB-231 cells were performed with each siRNA pair for 48 hours. The expression level of each gene of interest, relative to negative control siRNA transfections, was then determined by real-time RT-PCR in technical triplicates. All siRNA pairs knockdown the expression of the target gene by at least an average of 70 percent.

An effective and specific knockdown that minimizes false-negative and false-positive hit rates, respectively, is critical to the success of screening any siRNA library for phenotypic effects.