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Plant Gene Regulatory Networks pp 167–184Cite as

Multiplexed Transcriptional Activation or Repression in Plants Using CRISPR-dCas9-Based Systems

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Part of the book series: Methods in Molecular Biology ((MIMB,volume 1629))

Abstract

Novel tools and methods for regulating in vivo plant gene expression are quickly gaining popularity and utility due to recent advances in CRISPR-dCas9 chimeric effector regulators, otherwise known as CRISPR artificial transcription factors (CRISPR-ATFs). These tools are especially useful for studying gene function and interaction within various regulatory networks. First generation CRISPR-ATFs are nuclease-deactivated (dCas9) CRISPR systems where dCas9 proteins are fused to known transcriptional activator domains (VP64) or repressor domains (SRDX). When multiple chimeric dCas9-effector fusions are guided to gene regulatory regions via CRISPR gRNAs, they can modulate expression of transcript levels in planta. The protocol presented here provides a detailed procedure for activating AtPAP1 and repressing AtCSTF64 in Arabidopsis thaliana. This protocol makes use of our plant CRISPR toolbox to streamline the assembly and cloning of multiplex CRISPR-Cas9 transcriptional regulatory constructs.

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Acknowledgments

This work is supported by a Collaborative Funding Grant (2016-CFG-8003) from North Carolina Biotechnology Center and Syngenta Biotechnology to Y.Q.

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Authors and Affiliations

  1. Department of Biology, East Carolina University, Greenville, NC, 27858, USA

    Levi G. Lowder, Joseph W. Paul III & Yiping Qi

Authors
  1. Levi G. Lowder
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  2. Joseph W. Paul III
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  3. Yiping Qi
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Corresponding author

Correspondence to Yiping Qi .

Editor information

Editors and Affiliations

  1. Department for Plant Cell and Molecular Biology, Institute for Biology, Humboldt-Universität zu Berlin, Berlin, Germany

    Kerstin Kaufmann

  2. Institute for Biochemistry and Biology, University of Potsdam, Potsdam, Germany

    Bernd Mueller-Roeber

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Lowder, L.G., Paul, J.W., Qi, Y. (2017). Multiplexed Transcriptional Activation or Repression in Plants Using CRISPR-dCas9-Based Systems. In: Kaufmann, K., Mueller-Roeber, B. (eds) Plant Gene Regulatory Networks. Methods in Molecular Biology, vol 1629. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-7125-1_12

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  • DOI: https://doi.org/10.1007/978-1-4939-7125-1_12

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  • Publisher Name: Humana Press, New York, NY

  • Print ISBN: 978-1-4939-7124-4

  • Online ISBN: 978-1-4939-7125-1

  • eBook Packages: Springer Protocols

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