Design of active small interfering RNAs

Integrated DNA Technologies Inc, 1710 Commercial Park, Coralville, IA 52241, USA.
Current opinion in molecular therapeutics (Impact Factor: 3.42). 05/2007; 9(2):110-8.
Source: PubMed


Small interfering RNAs (siRNAs) have become the experimental tool of choice to suppress gene expression in a wide variety of organisms. Site selection and optimization does not appear to be as difficult for siRNAs as would be expected from historical experience with antisense oligonucleotides. Nevertheless, not all sites within a target gene perform equally. Significant progress has been made in defining sequence features that contribute to siRNA potency, and a variety of computational tools are available from academic and commercial sources to assist with siRNA design. Potential siRNA sequences should be screened for homology to other genes within the target organism's transcriptome to minimize cross-hybridization and inadvertent knockdown of unrelated genes via off-target effects. In addition to rational design criteria, chemical modification of the RNA can improve function by improving stability, reducing the potential for off-target effects and avoiding stimulation of the innate immune system.

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    • "RNAi function and is therefore counterproductive, a pattern of alternating 2'-O-methyl bases is comparable with the activity of unmodified RNA, and is quite stable in serum (Choung et al., 2006; Czauderna et al., 2003; Peek and Behlke, 2007) . Conjugation of peptides to the dsRNA is also developed, to increase cell membrane permeability (Ifediba et al., 2010). "
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    ABSTRACT: Molecular genetics insight into the pathogenesis of several neurodegenerative diseases, such as Alzheimer's disease, Parkinson's disease, Huntington's disease and amyotrophic lateral sclerosis, encourage direct interference with the activity of neurotoxic genes or the molecular activation of neuroprotective pathways. Oligonucleotide-based therapies are recently emerging as an efficient strategy for drug development and these can be employed as new treatments of neurodegenerative states. Here we review advances in this field in recent years which suggest an encouraging assessment that oligonucleotide technologies for targeting of RNAs will enable the development of new therapies and will contribute to preservation of brain integrity.
    Brain research 04/2014; 1584. DOI:10.1016/j.brainres.2014.04.005 · 2.84 Impact Factor
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    • "Moreover, the proposed method considers both desired and undesired nucleotide content for a specific sequence position. Widely used features such as thermodynamic (TD) features, N-Gram and N-GSK (general string kernel) features [21] and some coding features are studied individually and grouped altogether. We also focus on the 2 base-pair (bp) patterns to extract the sequence position specified feature (SPSF) and position coding (PC) features. "
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    ABSTRACT: Background RNA interference (RNAi) becomes an increasingly important and effective genetic tool to study the function of target genes by suppressing specific genes of interest. This system approach helps identify signaling pathways and cellular phase types by tracking intensity and/or morphological changes of cells. The traditional RNAi screening scheme, in which one siRNA is designed to knockdown one specific mRNA target, needs a large library of siRNAs and turns out to be time-consuming and expensive. Results In this paper, we propose a conceptual model, called compressed sensing RNAi (csRNAi), which employs a unique combination of group of small interfering RNAs (siRNAs) to knockdown a much larger size of genes. This strategy is based on the fact that one gene can be partially bound with several small interfering RNAs (siRNAs) and conversely, one siRNA can bind to a few genes with distinct binding affinity. This model constructs a multi-to-multi correspondence between siRNAs and their targets, with siRNAs much fewer than mRNA targets, compared with the conventional scheme. Mathematically this problem involves an underdetermined system of equations (linear or nonlinear), which is ill-posed in general. However, the recently developed compressed sensing (CS) theory can solve this problem. We present a mathematical model to describe the csRNAi system based on both CS theory and biological concerns. To build this model, we first search nucleotide motifs in a target gene set. Then we propose a machine learning based method to find the effective siRNAs with novel features, such as image features and speech features to describe an siRNA sequence. Numerical simulations show that we can reduce the siRNA library to one third of that in the conventional scheme. In addition, the features to describe siRNAs outperform the existing ones substantially. Conclusions This csRNAi system is very promising in saving both time and cost for large-scale RNAi screening experiments which may benefit the biological research with respect to cellular processes and pathways.
    BMC Bioinformatics 12/2012; 13(1):337. DOI:10.1186/1471-2105-13-337 · 2.58 Impact Factor
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    • "Furthermore, the siRNA-induced off-target effects [45] and immunostimulatory activities may differ depending on the selected sequence [39]. Although a variety of siRNA design tools have been developed, not all potentially risky motifs have been discovered [46] bringing additional complexity and unpredictability to the selection process. The difficult design procedure of single-site siRNAs can, however, be largely overcome by using pools of siRNAs, applicable as single-site siRNAs, for example in a topical fashion [29]. "
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    ABSTRACT: RNA interference (RNAi)-based sequence-specific gene silencing is applied to identify gene function and also possesses great potential for inhibiting virus replication both in animals and plants. Small interfering RNA (siRNA) molecules are the inducers of gene silencing in the RNAi pathway but may also display immunostimulatory activities and promote apoptosis. Canonical siRNAs are 21 nucleotides (nt) in length and are loaded to the RNA Induced Silencing Complex when introduced into the cells, while longer siRNA molecules are first processed by endogenous Dicer and thus termed Dicer-substrate siRNA (DsiRNA). We have applied RNA polymerases from bacteriophages T7 and phi6 to make high-quality double-stranded RNA molecules that are specific for the UL29 gene of herpes simplex virus (HSV). The 653 nt long double-stranded RNA molecules were converted to siRNA and DsiRNA pools using Dicer enzymes originating from human or Giardia intestinalis, producing siRNAs of approximately 21 and 27 nt in length, respectively. Chemically synthesised 21 and 27 nt single-site siRNA targeting the UL29 were used as references. The impact of these siRNAs on cell viability, inflammatory responses, gene silencing, and anti-HSV activity were assayed in cells derived from human nervous system and skin. Both pools and the canonical single-site siRNAs displayed substantial antiviral activity resulting in four orders of magnitude reduction in virus titer. Notably, the pool of DsiRNAs caused lower immunostimulation than the pool of canonical siRNAs, whereas the immunostimulation effect was in relation to the length with the single-site siRNAs. Our results also propose differences in the processivity of the two Dicers.
    PLoS ONE 11/2012; 7(11):e51019. DOI:10.1371/journal.pone.0051019 · 3.23 Impact Factor
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