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siRNA Design: It's All in the Algorithm

aiming to offer high success rates for silencing human genes. The purported performance of some of the resulting siRNA design algorithms has been based on largely statistical extrapolations and/or experimental short-cuts such as silencing of cotransfected exogenous reporter constructs. In collaboration with Ambion, Cenix BioScience has chosen the more rigorous path of directly measuring the effectiveness of approximately 1,100 of its own algorithm-designed siRNAs in silencing almost 400 endogenously expressed human transcripts. Importantly, the analysis was carried out in cultured human cells under stringently standardized experimental conditions. This empirical study therefore represents the most physiologically relevant, direct, and comprehensive performance analysis of any siRNA design algorithm available today.


Maximize Success Rate of siRNAs
Publicly available siRNA design programs have so far shown success rates of 50-60% in generating siRNAs that can yield over 70% silencing of target mRNA levels in HeLa cells after 48 hr. Based on this benchmark, Cenix conducted a first test of its algorithm's success rate by measuring silencing efficacy under the same conditions, namely using quantitative RT-PCR to measure target mRNA levels in HeLa cells 48 hr after transfection, for a set of 79 algorithm-designed siRNAs, each of which targeted a different kinase transcript. The analysis revealed that 74 of the 79 siRNAs, or 94% of the tested siRNAs, gave higher than 70% silencing (see Figure 1).

Figure 1. Silencing Efficacy of 79 siRNAs Designed Using Cenix Algorithm to Target 79 Endogenously-expressed Human Kinases. Target mRNA levels were meas
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