Fluorescent Guide RNAs Facilitate Development of Layered Pol II-Driven CRISPR Circuits

David J. Menn, Swechchha Pradhan, Samira Kiani, Xiao Wang

Research output: Contribution to journalArticlepeer-review

4 Scopus citations

Abstract

Efficient clustered regularly interspaced short palindromic repeat (CRISPR) guide RNA (gRNA) expression from RNA Polymerase II (Pol II) promoters will aid in construction of complex CRISPR-based synthetic gene networks. Yet, we require tools to properly visualize gRNA directly to quantitatively study the corresponding network behavior. To address this need, we employed a fluorescent gRNA (fgRNA) to visualize synthetic CRISPR network dynamics without affecting gRNA functionality. We show that studying gRNA dynamics directly enables circuit modification and improvement of network function in Pol II-driven CRISPR circuits. This approach generates information necessary for optimizing the overall function of these networks and provides insight into the hurdles remaining in Pol II-regulated gRNA expression. ©

Original languageEnglish (US)
Pages (from-to)1929-1936
Number of pages8
JournalACS Synthetic Biology
Volume7
Issue number8
DOIs
StatePublished - Aug 17 2018

Keywords

  • CRISPR
  • computational biology
  • fgRNA
  • genetic circuit
  • synthetic biology

ASJC Scopus subject areas

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

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