Apr 18, 2020

Membrane-bound GFP-labelled vectors for Targeted DamID allow simultaneous profiling of expression domains and DNA binding

BioRxiv : the Preprint Server for Biology
C. DelandreOwen J Marshall

Abstract

Targeted DamID (TaDa) allows highly efficient cell-type-specific profiling of protein-DNA interactions. Cell-type-specificity, however, is governed by the GAL4/UAS system, which can exhibit differences in expression patterns depending upon the genomic insertion site and the UAS promoter strength. The TaDa system uses a bicistronic transcript to reduce the translation rates of Dam-fusion proteins, presenting the possibility of using the primary ORF within in the transcript to label expression domains and precisely identified the profiled cell populations in experimental samples. Here, we describe two TaDa vectors, pTaDaG and pTaDaG2, that use myristoylated GFP as the primary ORF. Differing lengths of the myristoylation sequence between the plasmids allows additional translational control. Fly lines created with this system allow easy visualisation of expression domains under both fluorescent dissecting and confocal microscopes without the use of antibody staining, whilst faithfully profiling protein-DNA interactions via Targeted DamID.

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Mentioned in this Paper

Study
Flow Cytometry
HLA-DRB4
DNA Probes
Human Volunteers
Nucleic Acid Hybridization Procedure
SLC4A1 wt Allele
Fluorescence-Activated Cell Sorting
Keratin
HLA Antigens

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