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CRISPR / C as9‐mediated gene knockout in the ascidian C iona intestinalis
Author(s) -
Sasaki Haruka,
Yoshida Keita,
Hozumi Akiko,
Sasakura Yasunori
Publication year - 2014
Publication title -
development, growth and differentiation
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.864
H-Index - 66
eISSN - 1440-169X
pISSN - 0012-1592
DOI - 10.1111/dgd.12149
Subject(s) - crispr , rna , gene , biology , mutagenesis , ciona intestinalis , genetics , gene knockout , genome editing , guide rna , crispr interference , computational biology , mutation , microbiology and biotechnology
Knockout of genes with CRISPR / C as9 is a newly emerged approach to investigate functions of genes in various organisms. We demonstrate that CRISPR / C as9 can mutate endogenous genes of the ascidian C iona intestinalis , a splendid model for elucidating molecular mechanisms for constructing the chordate body plan. Short guide RNA (sg RNA ) and C as9 m RNA , when they are expressed in C iona embryos by means of microinjection or electroporation of their expression vectors, introduced mutations in the target genes. The specificity of target choice by sg RNA is relatively high compared to the reports from some other organisms, and a single nucleotide mutation at the sg RNA dramatically reduced mutation efficiency at the on‐target site. CRISPR / C as9‐mediated mutagenesis will be a powerful method to study gene functions in C iona along with another genome editing approach using TALE nucleases.