rmce Molecular mechanisms
Recombinase-Mediated Cassette Exchange (RMCE) is a targeted recombination technique that allows the exchange of a pre-existing genetic cassette in the Drosophila melanogaster genome with a transgene of interest. It relies on specific recombination sites (such as loxP for Cre or attP/attB for ΦC31), ensuring controlled and stable insertion of the transgene.
Microinjection Steps :
1. Anchoring Site Preparation :
A cassette containing the recombination sites is already integrated into the Drosophila genome (often via the MIMIC or ΦC31 system).
This cassette can contain a visible (white or yellow) or fluorescent marker.
2. Transgene and Recombinase Microinjection :
A plasmid DNA containing the gene of interest flanked by the same recombination sites is injected into early embryos.
A recombinase (Cre, ΦC31, or FLP) catalyzes the exchange between homologous sites, inserting the transgene at a defined locus.
3. Selection of modified individuals :
Flies are selected by phenotype change or molecular analysis (PCR).