Single vector non-leaky gene expression system for Drosophila melanogaster

Abstract

An ideal transgenic gene expression system is inducible, non-leaky, and well tolerated by the target organism. While the former has been satisfactorily realized, leakiness and heavy physiological burden imposed by the existing systems are still prominent hurdles in their successful implementation. Here we describe a new system for non-leaky expression of transgenes in Drosophila. PRExpress is based on a single transgenic construct built from endogenous components, the inducible hsp70 promoter and a multimerized copy of a Polycomb response element (PRE) controlled by epigenetic chromatin regulators of the Polycomb group. We show that this system is non-leaky, rapidly and strongly inducible, and reversible. To make the application of PRExpress user-friendly, we deliver the construct via site-specific integration.

A Drosophila CRISPR/Cas9 Toolkit for Conditionally Manipulating Gene Expression in the Prothoracic Gland as a Test Case for Polytene Tissues

“…In particular, Gal4 may result in signal amplification due to its strong activation domain, and one has only limited control over how strongly a given cDNA is expressed. Further, UAS-regulated transgenes all show some degree of leakiness, depending on the tissue and developmental time, potentially confounding experimental outcomes ( Akmammedov et al , 2017 ). Similarly, the presence of a second unrelated UAS-transgene may alter phenotypes seen with a single UAS-transgene alone, as both compete for Gal4-binding, which may quench the expression of either transgene.…”

A modified yeast three-hybrid system enabling both positive and negative selections

“…There was little to no inhibition at all 3-AT concentrations for the PUFwt expressing yeast. The lack of strong growth inhibition of the negative controls suggests that the HIS3 reporter in MaV204K is slightly leaky (hence the greater 3-AT concentration needed), this has been shown in similar HIS reporter systems in flies (Akmammedov et al, 2017).…”

Imaging cell lineage with a synthetic digital recording system

“…The resulting fly line, which we term ‘ Drosophila memoiphila ’, provides a resource for general purpose lineage analysis in flies. We further incorporated a Bxb1 integrase controlled by a tightly regulated heat shock inducible promoter ( 52 ) to record in specific time windows. Finally, we crossed these flies to an nSyb-Gal4 driver to restrict expression of the array to neurons (Fig.…”