Drosophila pp 155-174 | Cite as

Methods for Homologous Recombination in Drosophila

  • Keith A. Maggert
  • Wei J. Gong
  • Kent G. Golic
Part of the Methods in Molecular Biology book series (MIMB, volume 420)

Abstract

We present detailed protocols for two methods of gene targeting in Drosophila. The first, ends-out targeting, is identical in concept to gene replacement techniques used routinely in mammalian and yeast cells. In Drosophila, the targeted gene is replaced by the marker gene white+ (although options exist to generate unmarked targeted alleles). This approach is simple in both the molecular cloning and the genetic manipulations. Ends-out will likely serve most investigators’ purposes to generate simple gene deletions or reporter gene “knock-ins.”

The second method, ends-in targeting, targets a wild-type gene with an engineered mutated copy and generates a duplication structure at the target locus. This duplication can subsequently be reduced to one copy, removing the wild-type gene and leaving only the introduced mutation. Although more complicated in the cloning and genetic manipulations (seeNote 1), this approach has the benefit that the mutations may be introduced with no other remnant of the targeting procedure. This “surgical” approach will appeal to investigators who desire minimal perturbation to the genome, such as single nucleotide mutation.

Although both approaches appear to be approximately equally efficient (seeNote 2), each method has separate strengths and drawbacks. The choice of which approach is best depends on the researcher’s goal.

Key Words

Ends-in ends-out gene targeting homologous recombination mutation replacement 

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Copyright information

© Humana Press Inc., Totowa, NJ 2008

Authors and Affiliations

  • Keith A. Maggert
    • 1
  • Wei J. Gong
    • 2
  • Kent G. Golic
    • 2
  1. 1.Department of BiologyTexas A&M UniversityCollege StationUSA
  2. 2.Department of BiologyUniversity of UtahSalt Lake CityUSA

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