Design and Generation of Donor Constructs for Genome Engineering in Drosophila.

Citation:

Housden BE, Perrimon N. Design and Generation of Donor Constructs for Genome Engineering in Drosophila. Cold Spring Harb Protoc. 2016;2016 (9) :pdb.prot090787.

Date Published:

2016

Abstract:

The generation of precise alterations to the genome using CRISPR requires the combination of CRISPR and a donor construct containing homology to the target site. A double-strand break is first generated at the target locus using CRISPR. It is then repaired using the endogenous homologous recombination (HR) pathway. When a donor construct is provided, it can be used as a template for HR repair and can therefore be exploited to introduce alterations in the genomic sequence with single base-pair precision. Here we describe a protocol for the generation of donor constructs using Golden Gate assembly and discuss some key considerations for donor construct design for use in Drosophila.

See also: Review Article
Last updated on 06/12/2017