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Generating Mouse Models Using Zygote Electroporation of Nucleases (ZEN) Technology with High Efficiency and Throughput

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Zygotic Genome Activation

Part of the book series: Methods in Molecular Biology ((MIMB,volume 1605))

Abstract

Mouse models with genetic modifications are widely used in biology and biomedical research. Although the application of CRISPR-Cas9 system greatly accelerated the process of generating genetically modified mice, the delivery method depending on manual injection of the components into the embryos remains a bottleneck, as it is laborious, low throughput, and technically demanding. To overcome this limitation, we invented and optimized the ZEN (Zygote electroporation of nucleases) technology to deliver CRISPR-Cas9 reagents via electroporation. Using ZEN, we were able to generate genetically modified mouse models with high efficiency and throughput. Here, we describe the protocol in great detail.

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References

  1. Jinek M, Chylinski K, Fonfara I, Hauer M, Doudna JA, Charpentier E (2012) A programmable dual-RNA-guided DNA endonuclease in adaptive bacterial immunity. Science 337(6096):816–821. doi:10.1126/science.1225829. science.1225829 [pii]

    Article  CAS  PubMed  Google Scholar 

  2. Cong L, Ran FA, Cox D, Lin SL, Barretto R, Habib N, Hsu PD, Wu XB, Jiang WY, Marraffini LA, Zhang F (2013) Multiplex genome engineering using CRISPR/Cas systems. Science 339(6121):819–823. doi:10.1126/science.1231143

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  3. Mali P, Yang L, Esvelt KM, Aach J, Guell M, DiCarlo JE, Norville JE, Church GM (2013) RNA-guided human genome engineering via Cas9. Science 339(6121):823–826. doi:10.1126/science.1232033. science.1232033 [pii]

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  4. Wang H, Yang H, Shivalila CS, Dawlaty MM, Cheng AW, Zhang F, Jaenisch R (2013) One-step generation of mice carrying mutations in multiple genes by CRISPR/Cas-mediated genome engineering. Cell 153(4):910–918. doi:10.1016/j.cell.2013.04.025. S0092-8674(13)00467-4 [pii]

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  5. Yang H, Wang H, Shivalila CS, Cheng AW, Shi L, Jaenisch R (2013) One-step generation of mice carrying reporter and conditional alleles by CRISPR/Cas-mediated genome engineering. Cell 154(6):1370–1379. doi:10.1016/j.cell.2013.08.022. S0092-8674(13)01016-7 [pii]

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  6. Wang W, Kutny PM, Byers SL, Longstaff CJ, DaCosta MJ, Pang C, Zhang Y, Taft RA, Buaas FW, Wang H (2016) Delivery of Cas9 protein into mouse zygotes through a series of electroporation dramatically increases the efficiency of model creation. J Genet Genomics 43(5):319–327. doi:10.1016/j.jgg.2016.02.004. S1673-8527(16)30002-9 [pii]

    Article  PubMed  Google Scholar 

  7. Qin W, Dion SL, Kutny PM, Zhang Y, Cheng A, Jillette NL, Malhotra A, Geurts AM, Chen YG, Wang H (2015) Efficient CRISPR/Cas9-mediated genome editing in mice by zygote electroporation of nuclease. Genetics 200:423–430. doi:10.1534/genetics.115.176594. genetics.115.176594 [pii]

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  8. Chen S, Lee B, Lee AY, Modzelewski AJ, He L (2016) Highly efficient mouse genome editing by CRISPR ribonucleoprotein electroporation of zygotes. J Biol Chem 291(28):14457–14467. doi:10.1074/jbc.M116.733154. M116.733154 [pii]

    Article  CAS  PubMed  Google Scholar 

  9. Wiles MV, Qin W, Cheng AW, Wang H (2015) CRISPR-Cas9-mediated genome editing and guide RNA design. Mamm Genome 26(9–10):501–510. doi:10.1007/s00335–015-9565-z

    Google Scholar 

  10. Qin W, Kutny PM, Maser RS, Dion SL, Lamont JD, Zhang Y, Perry GA, Wang H (2016) Generating mouse models using CRISPR-Cas9-mediated genome editing. Curr Protoc Mouse Biol 6(1):39–66. doi:10.1002/9780470942390.mo150178

    Article  PubMed  PubMed Central  Google Scholar 

  11. Byers SL, Payson SJ, Taft RA (2006) Performance of ten inbred mouse strains following assisted reproductive technologies (ARTs). Theriogenology 65(9):1716–1726. doi:10.1016/j.theriogenology.2005.09.016. S0093-691X(05)00405-X [pii]

    Article  PubMed  Google Scholar 

  12. Brinkman EK, Chen T, Amendola M, van Steensel B (2014) Easy quantitative assessment of genome editing by sequence trace decomposition. Nucleic Acids Res 42(22):e168. doi:10.1093/nar/gku936. gku936 [pii]

    Article  PubMed  PubMed Central  Google Scholar 

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Author information

Authors and Affiliations

  1. The Jackson Laboratory, Bar Harbor, ME, 04609, USA

    Wenbo Wang, Yingfan Zhang & Haoyi Wang

  2. State Key Laboratory of Stem Cell and Reproductive Biology, Institute of Zoology, Chinese Academy of Sciences, Beijing, 100101, China

    Haoyi Wang

  3. The University of Pennsylvania Perelman School of Medicine, Philadelphia, PA, 19104-5156, USA

    Wenbo Wang

Authors
  1. Wenbo Wang
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  2. Yingfan Zhang
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  3. Haoyi Wang
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Corresponding author

Correspondence to Haoyi Wang .

Editor information

Editors and Affiliations

  1. Department of Animal and Poultry Sciences, Virginia Tech, Blacksburg, Virginia, USA

    Kiho Lee

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Wang, W., Zhang, Y., Wang, H. (2017). Generating Mouse Models Using Zygote Electroporation of Nucleases (ZEN) Technology with High Efficiency and Throughput. In: Lee, K. (eds) Zygotic Genome Activation. Methods in Molecular Biology, vol 1605. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-6988-3_15

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  • DOI: https://doi.org/10.1007/978-1-4939-6988-3_15

  • Published:

  • Publisher Name: Humana Press, New York, NY

  • Print ISBN: 978-1-4939-6986-9

  • Online ISBN: 978-1-4939-6988-3

  • eBook Packages: Springer Protocols

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