CRISPR/Cas9-mediated genome editing in sea urchins
2019
Abstract The
CRISPR(clustered regularly interspaced short
palindromicrepeat)/
Cas9(
CRISPR-associated nuclease 9) technology enables rapid, targeted, and efficient changes in the genomes of various model organisms. The short
guide RNAs(gRNAs) of the
CRISPR/
Cas9system can be designed to recognize target DNA within coding regions for functional
gene knockouts. Several studies have demonstrated that the
CRISPR/
Cas9system efficiently and specifically targets
sea urchingenes and results in expected mutant phenotypes. In addition to disrupting gene functions, modifications and additions to the
Cas9protein enable alternative activities targeted to specific sites within the genome. This includes a fusion of
cytidine deaminaseto
Cas9(
Cas9-DA) for single nucleotide conversion in targeted sites. In this chapter, we describe detailed methods for the
CRISPR/
Cas9application in
sea urchinembryos, including gRNA design, in vitro synthesis of single
guide RNA(sgRNA), and the usages of the
CRISPR/
Cas9technology for
gene knockoutand single nucleotide editing. Methods for genotyping the resultant embryos are also provided for assessing efficiencies of gene editing.
Keywords:
-
Correction
-
Source
-
Cite
-
Save
28
References
3
Citations
NaN
KQI