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Genome targeting by hybrid Flp-TAL recombinases
Eugenia Voziyanova1, Feng Li1, Riddhi Shah2
1School of Biological Sciences, Louisiana Tech University, 1 Adams Blvd., Ruston, LA, 71272, USA.
Scientific Reports
|October 16, 2020
Summary
We developed Flp-TAL recombinases, novel hybrid enzymes for precise genome engineering. These tools enable targeted DNA integration and deletion at specific genomic locations, advancing gene editing capabilities.
Area of Science:
- Molecular Biology
- Genetics
- Biotechnology
Background:
- Genome engineering relies on diverse tools for precise DNA manipulation.
- Site-specific recombinases like Flp are crucial for targeted genomic modifications.
Purpose of the Study:
- To develop and characterize novel Flp-TAL recombinases for targeting native genomic FRT-like sequences.
- To demonstrate the utility of Flp-TAL recombinases in genome engineering applications.
Main Methods:
- Construction of hybrid Flp-TAL recombinases combining Flp recombinase and TAL effector DNA-binding domains.
- Functional validation of Flp-TAL recombinases in human HEK-293 cells using integration and deletion assays.
- Targeting of FRT-like sequences in the β-globin locus for integration experiments.
Main Results:
- Flp-TAL recombinases successfully targeted native genomic FRT-like sequences.
- Demonstrated targeted DNA integration into the β-globin locus with ~0.1% efficiency.
- Achieved precise excision of ~15 kilobases of genomic DNA with ~20% efficiency.
Conclusions:
- Flp-TAL recombinases represent a new class of genome engineering tools with targeted DNA manipulation capabilities.
- These enzymes offer a versatile platform for both integrating and deleting DNA sequences at specific genomic sites.
- The demonstrated functionality in human cells highlights the potential of Flp-TAL recombinases for various gene editing applications.
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