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Manufacturing and Delivering Genome-Editing Proteins

  • Jia LiuEmail author
  • Ya-jun Liang
  • Pei-ling Ren
  • Thomas GajEmail author
Protocol
Part of the Methods in Molecular Biology book series (MIMB, volume 1867)

Abstract

Genome-editing technologies have revolutionized the biomedical sciences by providing researchers with the ability to quickly and efficiently modify genes. While programmable nucleases can be introduced into cells using a variety of techniques, their delivery as purified proteins is an effective approach for limiting off-target effects. Here, we describe step-by-step procedures for manufacturing and delivering genome-modifying proteins—including Cas9 ribonucleoproteins (RNPs) and TALE and zinc-finger nucleases—into mammalian cells. Protocols for combining Cas9 RNP with naturally recombinogenic adeno-associated virus (AAV) donor vectors for the seamless insertion of transgenes by homology-directed genome editing are also provided.

Key words

ZFNs TALENs CRISPR RNP Protein delivery Genome editing 

Notes

Acknowledgments

This work was supported by Natural Science Foundation of China (No. 31600686 to J.L.) and ShanghaiTech University (Startup fund to the Laboratory of ADC Chemistry).

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Copyright information

© Springer Science+Business Media, LLC, part of Springer Nature 2018

Authors and Affiliations

  1. 1.Shanghai Institute for Advanced Immunochemical Studies (SIAIS)ShanghaiTech UniversityShanghaiChina
  2. 2.Shanghai Advanced Research InstituteChinese Academy of ScienceShanghaiChina
  3. 3.University of Chinese Academy of SciencesBeijingChina
  4. 4.Department of BioengineeringUniversity of Illinois at Urbana-ChampaignUrbanaUSA

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