Two-step CRISPR-Cas9 protocol for transposable element deletion in D. melanogaster natural populations

Abstract

Summary: We present a protocol for generating a precise deletion, without altering the genetic background of the strain, of a transposable element (TE) in a natural population of Drosophila melanogaster using two steps of CRISPR-Cas9 homology-directed repair. We describe steps for replacing the TE by a fluorescent marker and for subsequent marker removal using single-guide RNAs, repair plasmids, and microinjection. We also detail steps for screening the deletion of the TE and generating a homozygous mutant strain.For complete details on the use and execution of this protocol, please refer to Merenciano and Gonzalez.1 : Publisher’s note: Undertaking any experimental protocol requires adherence to local institutional guidelines for laboratory safety and ethics.

Keywords

• CRISPR
• Evolutionary Biology
• Genetics
• Model Organisms
• Molecular Biology