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A Customizable Protocol for String Assembly gRNA Cloning (STAgR)
JoVE Journal
Genetics
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JoVE Journal Genetics
A Customizable Protocol for String Assembly gRNA Cloning (STAgR)

A Customizable Protocol for String Assembly gRNA Cloning (STAgR)

DOI:
10.3791/58556-v

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10:00 min

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December 26, 2018

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, , , , ,
1MCN Junior Research Group, Munich Center for Neurosciences,Ludwig Maximilian Universitat, BioMedical Center, 2Institute of Stem Cell Research, Helmholtz Zentrum,German Research Center for Environmental Health, 3Institute of Developmental Genetics, Helmholtz Zentrum,German Research Center for Environmental Health, 4Physiological Genomics,Ludwig Maximilian Universitat, BioMedical Center

Chapters

  • 00:04Title
  • 00:34Design of STAgR Cloning Primers with Overhangs
  • 01:22Generation of STAgR Cloning Fragments
  • 03:56Gibson Assembly Reaction and Bacterial Transformation
  • 05:50Selecting STAgR Clones by Bacterial Colony PCR
  • 08:08Results: Analysis of String Assembly gRNA Cloning
  • 09:27Conclusion

Summary

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Here, we present string assembly gRNA cloning (STAgR), a method to easily multiplex gRNA vectors for CRISPR/Cas9 approaches. STAgR makes gRNA multiplexing simple, efficient and customizable.

Tags

Keywords: CRISPR-Cas9GRNA CloningSTAgRMultiplex GRNAGibson AssemblyDNA TemplatePCRMagnetic BeadsDNA Purification

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