Use of the CRISPR-Cas9 system for genome editing in cultured Drosophila ovarian somatic cells

Hirotsugu Ishizu, Tetsutaro Sumiyoshi, Mikiko C. Siomi

Research output: Contribution to journalArticle

4 Citations (Scopus)

Abstract

The CRISPR-Cas9 system can be used for genome engineering in many organisms. PIWI-interacting RNAs (piRNAs) play a crucial role in repressing transposons to maintain genome integrity in Drosophila ovaries, and cultured ovarian somatic cells (OSCs) are widely used to elucidate the molecular mechanisms underlying the piRNA pathway. However, the germline-specific piRNA amplification system known as the ping-pong machinery does not occur in OSCs, making them unsuitable for elucidating the underlying mechanisms. Mutations in the lethal (3) malignant brain tumor gene (l(3)mbt) have been shown to cause ectopic expression of germline genes, including ping-pong factors. We therefore performed genome editing of Drosophila OSCs using the CRISPR-Cas9 system to achieve l(3)mbt knockout, resulting in successful induction of the piRNA amplification machinery. Here, we describe the detailed procedures for generating knockout and knockin OSC cells.

Original languageEnglish
Pages (from-to)186-192
Number of pages7
JournalMethods
Volume126
DOIs
Publication statusPublished - 2017 Aug 15
Externally publishedYes

Fingerprint

Clustered Regularly Interspaced Short Palindromic Repeats
Drosophila
Genes
Small Interfering RNA
Brain Neoplasms
Machinery
Amplification
Tumors
Genome
Brain
Gene Knockout Techniques
Ovary
Gene Editing
RNA
Mutation

ASJC Scopus subject areas

  • Molecular Biology
  • Biochemistry, Genetics and Molecular Biology(all)

Cite this

Use of the CRISPR-Cas9 system for genome editing in cultured Drosophila ovarian somatic cells. / Ishizu, Hirotsugu; Sumiyoshi, Tetsutaro; Siomi, Mikiko C.

In: Methods, Vol. 126, 15.08.2017, p. 186-192.

Research output: Contribution to journalArticle

Ishizu, Hirotsugu ; Sumiyoshi, Tetsutaro ; Siomi, Mikiko C. / Use of the CRISPR-Cas9 system for genome editing in cultured Drosophila ovarian somatic cells. In: Methods. 2017 ; Vol. 126. pp. 186-192.
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