Header

UZH-Logo

Maintenance Infos

Generation of genome-modified Drosophila cell lines using SwAP


Franz, Alexandra; Brunner, Erich; Basler, Konrad (2017). Generation of genome-modified Drosophila cell lines using SwAP. Fly:Epub ahead of print.

Abstract

The ease of generating genetically modified animals and cell lines has been markedly increased by the recent development of the versatile CRISPR/Cas9 tool. However, while the isolation of isogenic cell populations is usually straightforward for mammalian cell lines, the generation of clonal Drosophila cell lines has remained a longstanding challenge, hampered by the difficulty of getting Drosophila cells to grow at low densities. Here, we describe a highly efficient workflow to generate clonal Cas9-engineered Drosophila cell lines using a combination of cell pools, limiting dilution in conditioned medium and PCR with allele-specific primers, enabling the efficient selection of a clonal cell line with a suitable mutation profile. We validate the protocol by documenting the isolation, selection and verification of eight independently Cas9-edited armadillo mutant Drosophila cell lines. Our method provides a powerful and simple workflow that improves the utility of Drosophila cells for genetic studies with CRISPR/Cas9.

Abstract

The ease of generating genetically modified animals and cell lines has been markedly increased by the recent development of the versatile CRISPR/Cas9 tool. However, while the isolation of isogenic cell populations is usually straightforward for mammalian cell lines, the generation of clonal Drosophila cell lines has remained a longstanding challenge, hampered by the difficulty of getting Drosophila cells to grow at low densities. Here, we describe a highly efficient workflow to generate clonal Cas9-engineered Drosophila cell lines using a combination of cell pools, limiting dilution in conditioned medium and PCR with allele-specific primers, enabling the efficient selection of a clonal cell line with a suitable mutation profile. We validate the protocol by documenting the isolation, selection and verification of eight independently Cas9-edited armadillo mutant Drosophila cell lines. Our method provides a powerful and simple workflow that improves the utility of Drosophila cells for genetic studies with CRISPR/Cas9.

Statistics

Altmetrics

Downloads

4 downloads since deposited on 12 Sep 2017
4 downloads since 12 months
Detailed statistics

Additional indexing

Item Type:Journal Article, refereed, original work
Communities & Collections:07 Faculty of Science > Institute of Molecular Life Sciences
Dewey Decimal Classification:570 Life sciences; biology
Language:English
Date:30 August 2017
Deposited On:12 Sep 2017 15:13
Last Modified:12 Sep 2017 15:14
Publisher:Taylor & Francis
ISSN:1933-6934
Free access at:Publisher DOI. An embargo period may apply.
Publisher DOI:https://doi.org/10.1080/19336934.2017.1372068
PubMed ID:28853976

Download

Download PDF  'Generation of genome-modified Drosophila cell lines using SwAP'.
Preview
Content: Accepted Version
Language: English
Filetype: PDF
Size: 1MB
View at publisher