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논문검색

Poster 11

Cleavage of double-stranded DNA by engineered FokⅠendonuclease

초록

영어

Genome editing is one of the most demanding tools in mammalian cells to unravel the role of genes and to implement gene therapy of genetic diseases. Zinc finger nuclease is an engineered restriction enzyme which is constructed by combining an array of zinc finger DNA-binding motifs in the N- terminus and the FokⅠ endonuclease in C-terminus. Almost every ZFN recognizes three consecutive nucleotides starting with guanine in a context-dependent manner, thereby limiting the use of ZFN in genome editing. To address this challenge, we replaced ZFNs with a gene-specific nucleotides, which were connected to FokⅠ nuclease by using (his)6-tag and GS linkers. We tested in vitro whether this engineered FokⅠ shows a site-specific nuclease activity. For this, the engineered FokⅠ and nucleotide complex was incubated with template DNA which was produced from 70-mer and 100-mer nucleotides base-paired each other. The results indicate that the complex showed a site-specific nuclease activity, which suggests that the nucleotide-based FokⅠ can be used for the universal genome editing.

저자정보

  • Seong Kook Jeon Korean Research Institute of Bioscience & Biotechnology, Daejeon 305-806, Korea
  • Yong-Sam Kim Korean Research Institute of Bioscience & Biotechnology, Daejeon 305-806, Korea
  • Sun Hee Kim Korean Research Institute of Bioscience & Biotechnology, Daejeon 305-806, Korea
  • Jeong-Heon Ko Korean Research Institute of Bioscience & Biotechnology, Daejeon 305-806, Korea

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