Programmable DNA-Guided Artificial Restriction Enzymes

Behnam Enghiad, Huimin Zhao

Research output: Contribution to journalArticlepeer-review

Abstract

Restriction enzymes are essential tools for recombinant DNA technology that have revolutionized modern biological research. However, they have limited sequence specificity and availability. Here we report a Pyrococcus furiosus Argonaute (PfAgo) based platform for generating artificial restriction enzymes (AREs) capable of recognizing and cleaving DNA sequences at virtually any arbitrary site and generating defined sticky ends of varying length. Short DNA guides are used to direct PfAgo to target sites for cleavage at high temperatures (>87 °C) followed by reannealing of the cleaved single stranded DNAs. We used this platform to generate over 18 AREs for DNA fingerprinting and molecular cloning of PCR-amplified or genomic DNAs. These AREs work as efficiently as their naturally occurring counterparts, and some of them even do not have any naturally occurring counterparts, demonstrating easy programmability, generality, versatility, and high efficiency for this new technology.

Original languageEnglish (US)
Pages (from-to)752-757
Number of pages6
JournalACS synthetic biology
Volume6
Issue number5
DOIs
StatePublished - May 19 2017

Keywords

  • DNA cloning
  • DNA profiling
  • recombinant DNA technology
  • restriction enzymes

ASJC Scopus subject areas

  • Biomedical Engineering
  • Biochemistry, Genetics and Molecular Biology (miscellaneous)

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