CIRCLE-Seq for Interrogation of Off-Target Gene Editing.

Circularization for In Vitro Reporting of Cleavage Effects by Sequencing (CIRCLE-seq) is a novel technique developed for the impartial identification of unintended cleavage sites of CRISPR-Cas9 through targeted sequencing of CRISPR-Cas9 cleaved DNA. The protocol involves circularizing genomic DNA (gDNA), which is subsequently treated with the Cas9 protein and a guide RNA (gRNA) of interest. Following treatment, the cleaved DNA is purified and prepared as a library for Illumina sequencing.

Generation of an Induced Pluripotent Stem Cell Line with the Constitutive EGFP Reporter.

The discovery of induced pluripotent stem cell (iPSC) technology has provided a versatile platform for basic science research and regenerative medicine. With the rise of clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated protein 9 (Cas9) systems and the ease at which they can be utilized for gene editing, creating genetically modified iPSCs has never been more advantageous for studying both organism development and potential clinical applications. However, to better understand the behavior and true therapeutic potential of iPSCs and iPSC-derived cells, a tool for labeling and monitoring these cells in vitro and in vivo is needed.