siRNA design
siRNA design is the critical step of a successful RNAi experiment. Well designed siRNA sequence will minimize the off-targeting effect and maximize the knock down effect of the given gene. Most commonly cited siRNA design tools and algorithms are collected here, along with some siRNA design tools from major RNAi suppliers.
Published siRNA design algorithms and tools:
AsiDesigner for siRNA design
AsiDesigner: exon-based siRNA design server considering alternative splicing.
Park YK, Park SM, Choi YC, Lee D, Won M, Kim YJ.
AsiDesigner is a web-based siRNA design software system, which provides siRNA design capability to account for alternative splicing for mRNA level gene silencing. It provides numerous novel functions including the designing of common siRNAs for the silencing of more than two mRNAs simultaneously, a scoring scheme to evaluate the performance of designed siRNAs by adopting currently known key design factors, a stepwise off-target searching with BLAST and FASTA algorithms and checking the folding secondary structure energy of siRNAs.
DEQOR: a web-based tool for the design and quality control of siRNAs
Henschel A, Buchholz F, Habermann B.
Scionics Computer Innovation, GmbH, Pfotenhauerstrasse 110, 01307 Dresden, Germany.
DEQOR, uses a scoring system based on state-of-the-art parameters for siRNA design to evaluate the inhibitory potency of siRNAs. DEQOR, therefore, can help to predict (i) regions in a gene that show high silencing capacity based on the base pair composition and (ii) siRNAs with high silencing potential for chemical synthesis. In addition, each siRNA arising from the input query is evaluated for possible cross-silencing activities by performing BLAST searches against the transcriptome or genome of a selected organism. DEQOR can therefore predict the probability that an mRNA fragment will cross-react with other genes in the cell and helps researchers to design experiments to test the specificity of esiRNAs or chemically designed siRNAs.
siRNA design principles
- Length of 21 base pairs total
- Length of 19 base pairs
- A/U at the 5' anti-sense
- G/C at the 3' anti-sense
- Avoid G/C stretches (>7). 30-50% G/C overall.
- Limit <3 consecutive stretches of the same base
- No mismatches
- Thermodynamic constraints
- Hairpin formation
- Asymmetric ends
- 9-14 position of anti-sense has low stability
- Structural constraints
- target secondary structure
- siRNA secondary structure
- UTR and ATG may have binding factors
- Minimizing cross-hybridization with other nonspecific sequences
Computational siRNA resources and tools
Include (1) siRNA efficacy prediction: A Support vector regressor (SVR) for predicting efficacy for siRNAs; (2) siRNA specificity prediction: Specificity search for siRNAs to avoid off-target effects; (3) siRNAdb: A database of known siRNA molecules
siVirus: for efficient antiviral siRNA design.
Find highly conserved target sites; Design highly effective siRNAs; Design off-target minimized siRNAs. A web-based online software system that provides efficient short interfering RNA (siRNA) design for antiviral RNA interference (RNAi). siVirus searches for functional, off-target minimized siRNAs targeting highly conserved regions of divergent viral sequences.
Nucleic Acids Res. 2006 Jul 1;34(Web Server issue):W448-50.
siDRM: an effective and generally applicable online siRNA design tool
siDRM uses disjunctive rule merging (DRM) approach for siRNA design. A performance comparison using an independent dataset indicated that siDRM outperforms 19 existing siRNA design tools in identifying effective siRNAs.
Bioinformatics. 2008 Oct 15;24(20):2405-6. Epub 2008 Aug 20.
eRNAi
Genome-wide RNAi analysis of growth and viability in Drosophila cells.
Boutros M, Kiger AA, Armknecht S, Kerr K, Hild M, Koch B, Haas SA, Paro R, Perrimon N; Heidelberg Fly Array Consortium.
Science. 2004 Feb 6;303(5659):832-5.
E-RNAi is a tool for for designing siRNA for RNAi experiments in Drosophila and C. elegans. It can also be used for the design of enzymatically-digested long dsRNA (esiRNAs) for mammalian cells.
OligoWalk: an online siRNA design tool utilizing hybridization thermodynamics
Given an mRNA sequence as input, the OligoWalk web server generates a list of small interfering RNA (siRNA) candidate sequences, ranked by the probability of being efficient siRNA (silencing efficacy greater than 70%).
On average, the fraction of efficient siRNAs selected by the server that will be efficient at silencing is 78.6%.
Sfold: A bayesian statistical algorithm for RNA secondary structure prediction
sFold uses Bayesian algorithm that is based on stacking energy rules but relaxes the need to specify the parameters. The algorithm returns the exact posterior distribution of the number of destabilizing loops, stacking energy matrices, and secondary structures. The algorithm generates statistically representative structures from the full ensemble of probable secondary structures in exact proportion to the posterior probabilities.
siDirect
highly effective, target specific siRNA online design tool.
Guidelines for the selection of highly effective siRNA sequences for mammalian and chick RNA interference. Nucleic Acids Res. 2004 Feb 9;32(3):936-48.
siRNA Selection Server: an automated siRNA oligonucleotide prediction server
The Whitehead siRNA (short interfering RNA) Selection Web Server automates the design of short oligonucleotides that can specifically 'knock down' expression of target genes. These short sequences are about 21 nt in length, and when synthesized as double stranded RNA and introduced into cell culture, can reduce or eliminate the function of the target gene.
Commercial siRNA design tools:
siRNA target finder from Ambion
This tool employs the early siRNA design guidelines first described by Tuschl et.al. It has been found ~50% of siRNAs designed using this tool will reduce target gene expression by >50%.
siDESIGN Center from Dharmacon
Dharmacon offers this user-friendly siRNA design tool with detailed help information.
siRNA Target Finder from GenScript
Read siRNA design strategy before using this tool.
Free siRNA design tools:
- siRNA design tools and resources: siRNA design tools using support vector machine, shRNA oligo design and more.
- BLAST search tutorial from NCBI is essential for novice user to design their own siRNA sequences.
- BIOPREDsi siRNA design tool
- BIOPREDsi is an artificial intelligence algorithm that computationally predicts 21-nt siRNA sequences that have an optimal knockdown effect for a given gene. BIOPREDsi prediction algorithm is based on neuronal network.
Related content
Comments
Log in or create a user account to comment.
