Skip to main content
SpringerLink
Log in

Bioinformatics of siRNA Design

  • Protocol
  • First Online:
RNA Sequence, Structure, and Function: Computational and Bioinformatic Methods

Part of the book series: Methods in Molecular Biology ((MIMB,volume 1097))

Abstract

RNA interference mediated by small interfering RNAs is a powerful tool for investigation of gene functions and is increasingly used as a therapeutic agent. However, not all siRNAs are equally potent, and although simple rules for the selection of good siRNAs were proposed early on, siRNAs are still plagued with widely fluctuating efficiency. Recently, new design tools incorporating both the structural features of the targeted RNAs and the sequence features of the siRNAs substantially improved the efficacy of siRNAs. In this chapter we will present a review of sequence and structure-based algorithms behind them.

This is a preview of subscription content, log in via an institution to check access.

Access this chapter

Log in via an institution

Subscribe and save

Springer+ Basic
$34.99 /Month
  • Get 10 units per month
  • Download Article/Chapter or eBook
  • 1 Unit = 1 Article or 1 Chapter
  • Cancel anytime
Subscribe now

Buy Now

Protocol
USD 49.95
Price excludes VAT (USA)
  • Available as PDF
  • Read on any device
  • Instant download
  • Own it forever
eBook
USD 139.00
Price excludes VAT (USA)
  • Available as EPUB and PDF
  • Read on any device
  • Instant download
  • Own it forever
Softcover Book
USD 179.99
Price excludes VAT (USA)
  • Compact, lightweight edition
  • Dispatched in 3 to 5 business days
  • Free shipping worldwide - see info
Hardcover Book
USD 219.99
Price excludes VAT (USA)
  • Durable hardcover edition
  • Dispatched in 3 to 5 business days
  • Free shipping worldwide - see info

Tax calculation will be finalised at checkout

Purchases are for personal use only

Institutional subscriptions

Similar content being viewed by others

References

  1. Fire A, Xu S, Montgomery MK, Kostas SA, Driver SE, Mello CC (1998) Potent and specific genetic interference by double-stranded RNA in Caenorhabditis elegans. Nature 391(6669):806–811. http://www.hubmed.org/display.cgi?uids=9486653

    Google Scholar 

  2. Elbashir SM, Harborth J, Lendeckel W, Yalcin A, Weber K, Tuschl T (2001) Duplexes of 21-nucleotide RNAs mediate RNA interference in cultured mammalian cells. Nature 411(6836):494–498. http://www.hubmed.org/display.cgi?uids=11373684

    Google Scholar 

  3. Stein CA (2001) Antisense that comes naturally. Nat Biotechnol 19(8):737–738. doi:10.1038/90783. http://dx.doi.org/10.1038/90783

    Google Scholar 

  4. Holen T, Amarzguioui M, Wiiger MT, Babaie E, Prydz H (2002) Positional effects of short interfering RNAs targeting the human coagulation trigger Tissue Factor. Nucleic Acids Res 30(8):1757–1766. http://www.hubmed.org/display.cgi?uids=11937629

    Google Scholar 

  5. Patzel V (2007) In silico selection of active siRNA. Drug Discov Today 12(3–4):139–148. doi:10.1016/j.drudis.2006.11.015. http://dx.doi.org/10.1016/j.drudis.2006.11.015

  6. Elbashir SM, Martinez J, Patkaniowska A, Lendeckel W, Tuschl T (2001) Functional anatomy of siRNAs for mediating efficient RNAi in Drosophila melanogaster embryo lysate. EMBO J 20(23): 6877–6888. http://www.hubmed.org/display.cgi?uids=11726523

    Google Scholar 

  7. Khvorova A, Reynolds A, Jayasena SD (2003) Functional siRNAs and miRNAs exhibit strand bias. Cell 115(2):209–216. http://www.hubmed.org/display.cgi?uids=14567918

    Google Scholar 

  8. Schwarz DS, Hutvágner G, Du T, Xu Z, Aronin N, Zamore PD (2003) Asymmetry in the assembly of the RNAi enzyme complex. Cell 115(2):199–208

    Article  CAS  PubMed  Google Scholar 

  9. Amarzguioui M, Prydz H (2004) An algorithm for selection of functional siRNA sequences. Biochem Biophys Res Commun 316(4):1050–1058. http://www.hubmed.org/display.cgi?uids=15044091

    Google Scholar 

  10. Hohjoh H (2004) Enhancement of RNAi activity by improved siRNA duplexes. FEBS Lett 557(1–3):193–198. http://www.hubmed.org/display.cgi?uids=14741366

    Google Scholar 

  11. Hsieh AC, Bo R, Manola J, Vazquez F, Bare O, Khvorova A, Scaringe S, Sellers WR (2004) A library of siRNA duplexes targeting the phosphoinositide 3-kinase pathway: determinants of gene silencing for use in cell-based screens. Nucleic Acids Res 32(3):893–901. http://www.hubmed.org/display.cgi?uids=14769947

    Google Scholar 

  12. Takasaki S, Kotani S, Konagaya A (2004) An effective method for selecting siRNA target sequences in mammalian cells. Cell Cycle 3(6):790–795

    Article  CAS  PubMed  Google Scholar 

  13. Ui-Tei K, Naito Y, Takahashi F, Haraguchi T, Ohki-Hamazaki H, Juni A, Ueda R, Saigo K (2004) Guidelines for the selection of highly effective siRNA sequences for mammalian and chick RNA interference. Nucleic Acids Res 32(3):936–948. http://www.hubmed.org/display.cgi?uids=14769950

    Google Scholar 

  14. Reynolds A, Leake D, Boese Q, Scaringe S, Marshall WS, Khvorova A (2004) Rational siRNA design for RNA interference. Nat Biotechnol 22(3):326–330. http://www.hubmed.org/display.cgi?uids=14758366

    Google Scholar 

  15. Patzel V, Rutz S, Dietrich I, Köberle C, Scheffold A, Kaufmann SH (2005) Design of siRNAs producing unstructured guide-RNAs results in improved RNA interference efficiency. Nat Biotechnol 23(11):1440–1444. doi:10.1038/nbt1151. http://dx.doi.org/10.1038/nbt1151

    Google Scholar 

  16. Saetrom P (2004) Predicting the efficacy of short oligonucleotides in antisense and RNAi experiments with boosted genetic programming. Bioinformatics 20(17):3055–3063. http://www.hubmed.org/display.cgi?uids=15201190

    Google Scholar 

  17. Ren Y, Gong W, Xu Q, Zheng X, Lin D, Wang Y, Li T (2006) siRecords: an extensive database of mammalian siRNAs with efficacy ratings. Bioinformatics 22(8):1027–1028. doi:10.1093/bioinformatics/btl026. http://dx.doi.org/10.1093/bioinformatics/btl026

    Google Scholar 

  18. Huesken D, Lange J, Mickanin C, Weiler J, Asselbergs F, Warner J, Meloon B, Engel S, Rosenberg A, Cohen D, Labow M, Reinhardt M, Natt F, Hall J (2005) Design of a genome-wide siRNA library using an artificial neural network. Nat Biotechnol 23(8):995–1001. http://www.hubmed.org/display.cgi?uids=16025102

    Google Scholar 

  19. Lima WF, Monia BP, Ecker DJ, Freier SM (1992) Implication of RNA structure on antisense oligonucleotide hybridization kinetics. Biochemistry 31(48):12055–12061

    Article  CAS  PubMed  Google Scholar 

  20. Vickers TA, Wyatt JR, Freier SM (2000) Effects of RNA secondary structure on cellular antisense activity. Nucleic Acids Res 28(6):1340–1347. http://www.hubmed.org/display.cgi?uids=10684928

    Google Scholar 

  21. Mir KU, Southern EM (1999) Determining the influence of structure on hybridization using oligonucleotide arrays. Nat Biotechnol 17(8):788–792. doi:10.1038/11732. http://dx.doi.org/10.1038/11732

    Google Scholar 

  22. Milner N, Mir KU, Southern EM (1997) Selecting effective antisense reagents on combinatorial oligonucleotide arrays. Nat Biotechnol 15(6):537–541. doi:10.1038/nbt0697-537. http://dx.doi.org/10.1038/nbt0697-537

    Google Scholar 

  23. Zhao JJ, Lemke G (1998) Rules for ribozymes. Mol Cell Neurosci 11(1–2):92–97. doi:10.1006/mcne.1998.0669. http://www.hubmed.org/display.cgi?uids=9608536

    Google Scholar 

  24. Ding Y, Lawrence CE (2001) Statistical prediction of single-stranded regions in RNA secondary structure and application to predicting effective antisense target sites and beyond. Nucleic Acids Res 29(5):1034–1046

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  25. Bohula EA, Salisbury AJ, Sohail M, Playford MP, Riedemann J, Southern EM, Macaulay VM (2003) The efficacy of small interfering RNAs targeted to the type 1 insulin-like growth factor receptor (IGF1R) is influenced by secondary structure in the IGF1R transcript. J Biol Chem 278(18):15991–15997. http://www.hubmed.org/display.cgi?uids=12604614

    Google Scholar 

  26. Kretschmer-Kazemi Far R, Sczakiel G (2003) The activity of siRNA in mammalian cells is related to structural target accessibility: a comparison with antisense oligonucleotides. Nucleic Acids Res 31(15):4417–4424

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  27. Xu Y, Zhang H-Y, Thormeyer D, Larsson O, Du Q, Elmén J, Wahlestedt C, Liang Z (2003) Effective small interfering RNAs and phosphorothioate antisense DNAs have different preferences for target sites in the luciferase mRNAs. Biochem Biophys Res Commun 306(3):712–717

    Article  CAS  PubMed  Google Scholar 

  28. Vickers TA, Koo S, Bennett CF, Crooke ST, Dean NM, Baker BF (2003) Efficient reduction of target RNAs by small interfering RNA and RNase H-dependent antisense agents. A comparative analysis. J Biol Chem 278(9):7108–7118. http://www.hubmed.org/display.cgi?uids=12500975

    Google Scholar 

  29. Ding Y, Chan CY, Lawrence CE (2004) Sfold web server for statistical folding and rational design of nucleic acids. Nucleic Acids Res 32(Web Server issue):W135–W141. doi: 10.1093/nar/gkh449. http://dx.doi.org/10.1093/nar/gkh449

  30. Shao Y, Chan CY, Maliyekkel A, Lawrence CE, Roninson IB, Ding Y (2007) Effect of target secondary structure on RNAi efficiency. RNA 13(10):1631–1640. doi:10.1261/rna.546207. http://dx.doi.org/10.1261/rna.546207

    Google Scholar 

  31. Luo KQ, Chang DC (2004) The gene-silencing efficiency of siRNA is strongly dependent on the local structure of mRNA at the targeted region. Biochem Biophys Res Commun 318(1):303–310. http://www.hubmed.org/display.cgi?uids=15110788

    Google Scholar 

  32. Yoshinari K, Miyagishi M, Taira K (2004) Effects on RNAi of the tight structure, sequence and position of the targeted region. Nucleic Acids Res 32(2):691–699. http://www.hubmed.org/display.cgi?uids=14762201

    Google Scholar 

  33. Overhoff M, Alken M, Far RK, Lemaitre M, Lebleu B, Sczakiel G, Robbins I (2005) Local RNA target structure influences siRNA efficacy: a systematic global analysis. J Mol Biol 348(4):871–881. http://www.hubmed.org/display.cgi?uids=15843019

    Google Scholar 

  34. Schubert S, Grünweller A, Erdmann VA, Kurreck J (2005) Local RNA target structure influences siRNA efficacy: systematic analysis of intentionally designed binding regions. J Mol Biol 348(4):883–893. http://www.hubmed.org/display.cgi?uids=15843020

    Google Scholar 

  35. Brown JR Sanseau P (2005) A computational view of microRNAs and their targets. Drug Discov Today 10(8):595–601. http://www.hubmed.org/display.cgi?uids=15837603

    Google Scholar 

  36. Ameres SL, Martinez J, Schroeder R (2007) Molecular basis for target RNA recognition and cleavage by human RISC. Cell 130(1):101–112. doi:10.1016/j.cell.2007.04.037. http://dx.doi.org/10.1016/j.cell.2007.04.037

    Google Scholar 

  37. Lu ZJ, Mathews DH (2008) Oligowalk: an online siRNA design tool utilizing hybridization thermodynamics. Nucleic Acids Res 36(Web Server issue):W104–W108. doi:10.1093/nar/gkn250. http://dx.doi.org/10.1093/nar/gkn250

  38. Lu ZJ, Mathews DH (2008) Efficient siRNA selection using hybridization thermodynamics. Nucleic Acids Res 36(2):640–647. doi:10.1093/nar/gkm920. http://dx.doi.org/10.1093/nar/gkm920

    Google Scholar 

  39. Tafer H, Ameres SL, Obernosterer G, Gebeshuber CA, Schroeder R, Martinez J, Hofacker IL (2008) The impact of target site accessibility on the design of effective siRNAs. Nat Biotechnol 26(5):578–583. doi:10.1038/nbt1404. http://dx.doi.org/10.1038/nbt1404

    Google Scholar 

  40. Boese Q, Leake D, Reynolds A, Read S, Scaringe SA, Marshall WS, Khvorova A (2005) Mechanistic insights aid computational short interfering RNA design. Methods Enzymol 392:73–96. doi:10.1016/S0076-6879(04)92005-8. http://dx.doi.org/10.1016/S0076-6879(04)92005-8

    Google Scholar 

  41. Mückstein U, Tafer H, Hackermüller J, Bernhart SH, Stadler PF, Hofacker IL (2006) Thermodynamics of RNA-RNA binding. Bioinformatics 22(10):1177–1182. doi:10.1093/bioinformatics/btl024. http://www.hubmed.org/display.cgi?uids=16446276

    Google Scholar 

  42. Mückstein U, Tafer H, Bernhard SH, Hernandez-Rosales M, Vogel J, Stadler PF, Hofacker IL (2008) Translational control by RNA-RNA interaction: improved computation of RNA-RNA binding thermodynamics. In: Elloumi M, Küng J, Linial M, Murphy R, Schneider K, Toma C (eds) Bioinformatics research and development. Communications in computer and information science, vol 13. Springer, Berlin, pp 114–127. doi:10.1007/978-3-540-70600-7_9

    Chapter  Google Scholar 

  43. Hornung V, Guenthner-Biller M, Bourquin C, Ablasser A, Schlee M, Uematsu S, Noronha A, Manoharan M, Akira S, de Fougerolles A, Endres S, Hartmann G (2005) Sequence-specific potent induction of IFN-alpha by short interfering RNA in plasmacytoid dendritic cells through TLR7. Nat Med 11(3):263–270. doi:10.1038/nm1191. http://dx.doi.org/10.1038/nm1191

    Google Scholar 

  44. de Haro C, Méndez R, Santoyo J (1996) The eIF-2alpha kinases and the control of protein synthesis. FASEB J 10(12): 1378–1387

    PubMed  Google Scholar 

  45. Marques JT, Williams BRG (2005) Activation of the mammalian immune system by siRNAs. Nat Biotechnol 23(11):1399–1405. doi:10.1038/nbt1161. http://dx.doi.org/10.1038/nbt1161

    Google Scholar 

  46. Shao XD, Wu KC, Guo XZ, Xie M-J, Zhang J, Fan D-M (2008) Expression and significance of HERG protein in gastric cancer. Cancer Biol Ther 7(1):45–50

    Article  CAS  PubMed  Google Scholar 

  47. Saetrom P, Snove O (2004) A comparison of siRNA efficacy predictors. Biochem Biophys Res Commun 321(1):247–253. http://www.hubmed.org/display.cgi?uids=15358242

    Google Scholar 

  48. Vert JP, Foveau N, Lajaunie C, Vandenbrouck Y (2006) An accurate and interpretable model for siRNA efficacy prediction. BMC Bioinformatics

    Google Scholar 

  49. Matveeva O, Nechipurenko Y, Rossi L, Moore B, Saetrom P, Ogurtsov AY, Atkins JF, Shabalina SA (2007) Comparison of approaches for rational sirna design leading to a new efficient and transparent method. Nucleic Acids Res

    Google Scholar 

  50. Ding Y, Lawrence CE (2003) A statistical sampling algorithm for RNA secondary structure prediction. Nucleic Acids Res 31(24):7280–7301

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  51. Harborth J, Elbashir SM, Vandenburgh K, Manninga H, Scaringe SA, Weber K, Tuschl T (2003) Sequence, chemical, and structural variation of small interfering RNAs and short hairpin RNAs and the effect on mammalian gene silencing. Antisense Nucleic Acid Drug Dev 13(2):83–105. doi:10.1089/108729003321629638. http://dx.doi.org/10.1089/108729003321629638

  52. Shabalina SA, Spiridonov AN, Ogurtsov AY (2006) Computational models with thermodynamic and composition features improve siRNA design. BMC Bioinformatics 7:65. doi:10.1186/1471-2105-7-65. http://dx.doi.org/10.1186/1471-2105-7-65

    Google Scholar 

  53. Bernhart SH, Tafer H, Mückstein U, Flamm C, Stadler PF, Hofacker IL (2006) Partition function and base pairing probabilities of RNA heterodimers. Algorithms Mol Biol 1(1):3. doi:10.1186/1748-7188-1-3. http://www.hubmed.org/display.cgi?uids=16722605

    Google Scholar 

  54. Bompfünewerer AF, Backofen R, Bernhart SH, Hertel J, Hofacker IL, Stadler PF, Will S (2008) Variations on RNA folding and alignment: lessons from benasque. J Math Biol 56:119–144. doi:10.1007/s00285-007-0107-5

    Google Scholar 

  55. Haley B, Zamore PD (2004) Kinetic analysis of the RNAi enzyme complex. Nat Struct Mol Biol 11(7):599–606. http://www.hubmed.org/display.cgi?uids=15170178

    Google Scholar 

  56. Jackson AL, Bartz SR, Schelter J, Kobayashi SV, Burchard J, Mao M, Li B, Cavet G, Linsley PS ( 2003) Expression profiling reveals off-target gene regulation by RNAi. Nat Biotechnol 21(6):635–637. http://www.hubmed.org/display.cgi?uids=12754523

    Google Scholar 

Download references

Author information

Authors and Affiliations

  1. Institut fur Informatik, Universitat Leipzig, Leipzig, Germany

    Hakim Tafer

Authors
  1. Hakim Tafer
    View author publications

    You can also search for this author in PubMed Google Scholar

Editor information

Editors and Affiliations

  1. Center for non-coding RNA in Technology and Health, IKVH University of Copenhagen, Frederiksberg, Denmark

    Jan Gorodkin

  2. University of Washington Dept. Computer Science & Engineering, Seattle, Washington, USA

    Walter L. Ruzzo

Rights and permissions

Reprints and permissions

Copyright information

© 2014 Springer Science+Business Media New York

About this protocol

Cite this protocol

Tafer, H. (2014). Bioinformatics of siRNA Design. In: Gorodkin, J., Ruzzo, W. (eds) RNA Sequence, Structure, and Function: Computational and Bioinformatic Methods. Methods in Molecular Biology, vol 1097. Humana Press, Totowa, NJ. https://doi.org/10.1007/978-1-62703-709-9_22

Download citation

  • DOI: https://doi.org/10.1007/978-1-62703-709-9_22

  • Published:

  • Publisher Name: Humana Press, Totowa, NJ

  • Print ISBN: 978-1-62703-708-2

  • Online ISBN: 978-1-62703-709-9

  • eBook Packages: Springer Protocols

Publish with us

Policies and ethics

Search

Navigation