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CRISPR pp 77–89Cite as

Computational Detection of CRISPR/crRNA Targets

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Part of the Methods in Molecular Biology book series (MIMB,volume 1311)

Abstract

The CRISPR-Cas systems in bacteria and archaea provide protection by targeting foreign nucleic acids. The sequence of the “spacers” within CRISPR arrays specifically determines the targets in invader genomes. These spacers provide the short specific RNA nucleotide sequences within the guide crRNAs. In addition to complementarity in the spacer–target (protospacer) interaction, short flanking protospacer adjacent motifs (PAMs), or mismatching flanks have a discriminatory role in accurate target detection. Here, we describe a bioinformatic method, called CRISPRTarget, to use the sequence of a CRISPR array (e.g., predicted via CRISPRDetect/CRISPRDirection) to identify the foreign nucleic acids it targets.

Key words

  • CRISPR-Cas
  • RNAi
  • Protospacer adjacent motifs
  • crRNA
  • Noncoding RNA

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  • DOI: 10.1007/978-1-4939-2687-9_5
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References

  1. Richter C, Chang JT, Fineran PC (2012) Function and regulation of clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR associated (Cas) systems. Viruses 4:2291–2311

    CrossRef  PubMed Central  CAS  PubMed  Google Scholar 

  2. Westra ER, Buckling A, Fineran PC (2014) CRISPR-Cas systems: beyond adaptive immunity. Nat Rev Microbiol 12:317–326

    CrossRef  CAS  PubMed  Google Scholar 

  3. Drevet C, Pourcel C (2012) How to identify CRISPRs in sequencing data. Methods Mol Biol 905:15–27

    CAS  PubMed  Google Scholar 

  4. Lange SJ, Alkhnbashi OS, Rose D, Will S, Backofen R (2013) CRISPRmap: an automated classification of repeat conservation in prokaryotic adaptive immune systems. Nucleic Acids Res 41:8034–8044

    CrossRef  PubMed Central  CAS  PubMed  Google Scholar 

  5. Biswas A, Fineran PC, Brown CM (2014) Accurate computational prediction of the transcribed strand of CRISPR non-coding RNAs. Bioinformatics 30:1805–1813

    CrossRef  CAS  PubMed  Google Scholar 

  6. Biswas A, Gagnon JN, Brouns SJ, Fineran PC, Brown CM (2013) CRISPRTarget: bioinformatic prediction and analysis of crRNA targets. RNA Biol 10:817–827

    CrossRef  PubMed Central  CAS  PubMed  Google Scholar 

  7. Fineran PC, Gerritzen MJ, Suarez-Diez M, Kunne T, Boekhorst J, van Hijum SA, Staals RH, Brouns SJ (2014) Degenerate target sites mediate rapid primed CRISPR adaptation. Proc Natl Acad Sci U S A 111:E1629–E1638

    CrossRef  PubMed Central  CAS  PubMed  Google Scholar 

  8. Richter C, Dy RL, McKenzie RE, Watson BN, Taylor C, Chang JT, McNeil M, Staals RHJ, Fineran PC (2014) Priming in the Type I-F CRISPR-Cas system triggers strand-independent spacer acquisition, bi-directionally from the primed protospacer. Nucleic Acids Res 42(13):8516–8526

    CrossRef  PubMed Central  CAS  PubMed  Google Scholar 

  9. Barrangou R, Fremaux C, Deveau H, Richards M, Boyaval P, Moineau S, Romero DA, Horvath P (2007) CRISPR provides acquired resistance against viruses in prokaryotes. Science 315:1709–1712

    CrossRef  CAS  PubMed  Google Scholar 

  10. Deveau H, Barrangou R, Garneau JE, Labonte J, Fremaux C, Boyaval P, Romero DA, Horvath P, Moineau S (2008) Phage response to CRISPR-encoded resistance in Streptococcus thermophilus. J Bacteriol 190:1390–1400

    CrossRef  PubMed Central  CAS  PubMed  Google Scholar 

  11. Ran FA, Hsu PD, Wright J, Agarwala V, Scott DA, Zhang F (2013) Genome engineering using the CRISPR-Cas9 system. Nat Protoc 8:2281–2308

    CrossRef  PubMed Central  CAS  PubMed  Google Scholar 

  12. Fineran PC, Dy RL (2014) Gene regulation by engineered CRISPR-Cas systems. Curr Opin Microbiol 18:83–89

    CrossRef  CAS  PubMed  Google Scholar 

  13. Yang L, Mali P, Kim-Kiselak C, Church G (2014) CRISPR-Cas-mediated targeted genome editing in human cells. Methods Mol Biol 1114:245–267

    CrossRef  CAS  PubMed  Google Scholar 

  14. Bland C, Ramsey TL, Sabree F, Lowe M, Brown K, Kyrpides NC, Hugenholtz P (2007) CRISPR recognition tool (CRT): a tool for automatic detection of clustered regularly interspaced palindromic repeats. BMC Bioinformatics 8:209

    CrossRef  PubMed Central  PubMed  Google Scholar 

  15. Edgar RC (2007) PILER-CR: fast and accurate identification of CRISPR repeats. BMC Bioinformatics 8:18

    CrossRef  PubMed Central  PubMed  Google Scholar 

  16. Grissa I, Vergnaud G, Pourcel C (2007) CRISPRFinder: a web tool to identify clustered regularly interspaced short palindromic repeats. Nucleic Acids Res 35:W52–W57

    CrossRef  PubMed Central  PubMed  Google Scholar 

  17. Rousseau C, Gonnet M, Le Romancer M, Nicolas J (2009) CRISPI: a CRISPR interactive database. Bioinformatics 25:3317–3318

    CrossRef  PubMed Central  CAS  PubMed  Google Scholar 

  18. Skennerton CT, Imelfort M, Tyson GW (2013) Crass: identification and reconstruction of CRISPR from unassembled metagenomic data. Nucleic Acids Res 41:e105

    CrossRef  PubMed Central  CAS  PubMed  Google Scholar 

  19. Rho M, Wu YW, Tang H, Doak TG, Ye Y (2012) Diverse CRISPRs evolving in human microbiomes. PLoS Genet 8:e1002441

    CrossRef  PubMed Central  CAS  PubMed  Google Scholar 

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Acknowledgements

This work was supported by a Rutherford Discovery Fellowship from the Royal Society of NZ to PCF, by a Human Frontier Science Program Grant to Ian Macara, Anne Spang and CMB. AB was a recipient of a University of Otago Postgraduate Scholarship and a Postgraduate Publishing Bursary.

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Authors and Affiliations

  1. Department of Biochemistry, University of Otago, 56, Dunedin, 9054, New Zealand

    Ambarish Biswas & Chris M. Brown

  2. Department of Microbiology and Immunology, University of Otago, 56, Dunedin, 9054, New Zealand

    Peter C. Fineran

Authors
  1. Ambarish Biswas
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  2. Peter C. Fineran
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  3. Chris M. Brown
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Corresponding author

Correspondence to Chris M. Brown .

Editor information

Editors and Affiliations

  1. Department of Cell and Molecular Biology, Uppsala University, Uppsala, Sweden

    Magnus Lundgren

  2. Department of Molecular Biology MIMS, Umeå University, Umeå, Sweden

    Emmanuelle Charpentier

  3. Department of Microbiology and Immunology, University of Otago, Dunedin, New Zealand

    Peter C. Fineran

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Biswas, A., Fineran, P.C., Brown, C.M. (2015). Computational Detection of CRISPR/crRNA Targets. In: Lundgren, M., Charpentier, E., Fineran, P. (eds) CRISPR. Methods in Molecular Biology, vol 1311. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-2687-9_5

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  • DOI: https://doi.org/10.1007/978-1-4939-2687-9_5

  • Publisher Name: Humana Press, New York, NY

  • Print ISBN: 978-1-4939-2686-2

  • Online ISBN: 978-1-4939-2687-9

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