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STAR: A Simple TAL Effector Assembly Reaction Using Isothermal Assembly.

Sabine Gogolok1, Ute Köber1, Steven M Pollard2

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|May 14, 2018
PubMed
Summary
This summary is machine-generated.

This study introduces a simple TALE assembly reaction (STAR) for efficiently creating transcription activator-like effectors (TALEs). STAR enables rapid, manual production of numerous TALE DNA-binding domains for genetic manipulation, bypassing complex plasmid construction.

Keywords:
Gibson assemblyMammalianPlasmidTAL effectorTALEN

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Area of Science:

  • Molecular Biology
  • Synthetic Biology
  • Genetics

Background:

  • Transcription activator-like effectors (TALEs) possess programmable DNA-binding domains crucial for genetic manipulation.
  • Constructing plasmids for TALE DNA-binding domains is challenging due to repetitive sequences.

Purpose of the Study:

  • To develop a simplified method for assembling TALE DNA-binding domains.
  • To enable efficient generation of TALEs for targeting specific DNA sequences.

Main Methods:

  • A simple TALE assembly reaction (STAR) utilizing a 68-part plasmid library.
  • Protocol designed for manual production of TALEs targeting 17 bp sequences.

Main Results:

  • Successful creation of TALEs binding to 17 bp target sequences.
  • Manual production of tens of TALEs achieved within an 8-hour protocol.
  • Full-length, sequence-verified plasmids obtained within a few days.

Conclusions:

  • The STAR method offers a convenient and efficient approach for generating numerous TALENs or TALE-TFs.
  • Eliminates the need for extensive plasmid libraries and costly liquid handling equipment.