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FLASH assembly of TALENs for high-throughput genome editing.

Nature biotechnology | May 18, 2012

http://www.ncbi.nlm.nih.gov/pubmed/22484455

Engineered transcription activator–like effector nucleases (TALENs) have shown promise as facile and broadly applicable genome editing tools. However, no publicly available high-throughput method for constructing TALENs has been published, and large-scale assessments of the success rate and targeting range of the technology remain lacking. Here we describe the fast ligation-based automatable solid-phase high-throughput (FLASH) system, a rapid and cost-effective method for large-scale assembly of TALENs. We tested 48 FLASH-assembled TALEN pairs in a human cell–based EGFP reporter system and found that all 48 possessed efficient gene-modification activities. We also used FLASH to assemble TALENs for 96 endogenous human genes implicated in cancer and/or epigenetic regulation and found that 84 pairs were able to efficiently introduce targeted alterations. Our results establish the robustness of TALEN technology and demonstrate that FLASH facilitates high-throughput genome editing at a scale not currently possible with other genome modification technologies.

Pubmed ID: 22484455 RIS Download

Mesh terms: Automation | Base Sequence | Biotinylation | Computational Biology | DNA Restriction Enzymes | Endonucleases | Epigenesis, Genetic | Gene Targeting | Genetic Techniques | Genome, Human | Green Fluorescent Proteins | Humans | Molecular Sequence Data | Mutation | Neoplasms | Sequence Analysis, DNA | Sequence Homology, Nucleic Acid | Transcription, Genetic

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Associated grants

  • Agency: NIGMS NIH HHS, Id: DP1 GM105378
  • Agency: NIH HHS, Id: DP1 OD006862
  • Agency: NIH HHS, Id: DP1 OD006862
  • Agency: NIH HHS, Id: DP1 OD006862-01
  • Agency: NIH HHS, Id: DP1 OD006862-02
  • Agency: NHGRI NIH HHS, Id: P50 HG005550
  • Agency: NHGRI NIH HHS, Id: P50 HG005550-01
  • Agency: NHGRI NIH HHS, Id: P50 HG005550-02
  • Agency: NHGRI NIH HHS, Id: P50HG005550
  • Agency: NCI NIH HHS, Id: T32 CA009216
  • Agency: NCI NIH HHS, Id: T32 CA009216
  • Agency: NCI NIH HHS, Id: T32 CA009216-30
  • Agency: NCI NIH HHS, Id: T32 CA009216-31

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