CRISPR-Cas9系统在裂殖酵母中的应用

Jacobs Jake Z, Ciccaglione Keith M, Tournier Vincent, Zaratiegui Mikel

Department of Molecular Biology and Biochemistry, Rutgers, the State University of New Jersey, Piscataway, New Jersey 08854, USA.

Nat Commun. 2014 Oct 29;5:5344. doi: 10.1038/ncomms6344.

Application of the CRISPR-Cas9 genome editing system in the model organism Schizosaccharomyces pombe has been hampered by the lack of constructs to express RNA of arbitrary sequence. Here we present expression constructs that use the promoter/leader RNA of K RNA (rrk1) and a ribozyme to produce the targeting guide RNA. Together with constitutive expression of Cas9, this system achieves selection-free specific mutagenesis with efficiencies approaching 100%. The rrk1 CRISPR-Cas9 method enables rapid and efficient genome manipulation and unlocks the CRISPR toolset for use in fission yeast.

由于缺乏表达任意序列RNA的构建体，CRISPR-Cas9基因组编辑系统在模式生物粟酒裂殖酵母中的应用受到了阻碍。在此，我们展示了利用K RNA（rrk1）的启动子/前导RNA和一种核酶来产生靶向引导RNA的表达构建体。与Cas9的组成型表达一起，该系统实现了无选择的特异性诱变，效率接近100%。rrk1 CRISPR-Cas9方法能够实现快速高效的基因组操作，并为裂殖酵母解锁了CRISPR工具集。