Gal4 driver lines drosophila genetics

Uas gal4 system over expression tool in drosophila. Rnai fly lines based on the binary gal4 uas system are available commercially from several centers including vienna drosophila rnai center vdrc, the nigfly national institutes of genetics, japan and the trip transgenic rnai project collection at harvard medical school. Took an introductory genetics class at my university just a month ago and im looking at my notes on it so bear with me. Elav gal4 was used for constitutive expression of uastransgenes in. Oct 25, 2012 we established a collection of 7,000 transgenic lines of drosophila melanogaster. The authors describe the expression patterns driven by each fragment in the adult brain and ventral nerve cord and illustrate the utility of these data in identifying novel.

With geneswitch gal4 the expression of upstream activating sequence uas effector lines is controlled by. All gal4 insertions at bloomington known to be useful drivers of uasmediated expression are listed here. To facilitate genetic studies of reciprocal signaling, we have also made a plasmatocyteexpressing qf2 line that, in combination with extant gal4 drivers, allows independent genetic manipulation of both plasmatocytes and surrounding tissues, and gal80 lines that block gal4 drivers. The driver lines provide tissuespecific gal4 expression and the responder lines carry. Tissuespecific or ubiquitous enhancers drive expression of gal4 that results in transcriptional activation from the gal4 binding sites uas to drive. Third, we performed a genetic epistasis experiment by combining the expression of a constitutively active mutant rac1ca and the scarrnai1 in the mb neurons. A gal4 driver line resource for drosophila neurobiology. Rnai fly lines based on the binary gal4 uas system are available commercially from several centers including vienna drosophila rnai center vdrc, the nigfly national institutes of genetics, japan. Gal4 is then only expressed in cells where the driver gene is usually active. Three gal4 driver lines were used for the rnai crosses. Gal4 lines are one part of the binary gal4uas system brand and perrimon, 1993.

Therefore, analysis using these multiple binary expression systems has great potential for studying not only neuronglia but also gliaglia. The standard expression system used in drosophila is the bipartite gal4uas system first described by brand and perrimon 1993. In drosophila melanogaster, a widelyused technique for refined cell typespecific manipulation is the split gal4 system, which augments the targeting specificity of the binary gal4 uas upstream activating sequence system by making gal4 transcriptional activity contingent upon two enhancers, rather than one. A broad expression profile of the gmrgal4 driver in. Here, we report that the slbo gal4 transgene present in the line. The presence of gal4 in an alternating segmental pattern in the depicted embryos then drives expression of the uas responder gene in a corresponding. For example, overexpression of two genes by one gal4 driver will produce. In this report, we describe a drosophila screening tool, which we name the g al4. In order to label, monitor and manipulate a specific subtype of glial cells in a particular developmental or functional state with the gal4 uas system, it is necessary to obtain various gal4 lines that selectively express gal4 in each glial cell of interest.

The collection of gal4 driver lines we describe here should be particularly useful for identifying and characterizing the morphology of neuronal populations in the adult fly for several reasons. The data presented on this site are the work of the janelia flylight project team and the laboratories of gerald m. Most studies on bc migration have exploited a slbo gal4 driver to regulate gene expression in these cells or to mark them. Although rubin had little experience in either area hed spent his career studying molecular genetics and genomics in drosophila he knew that neuroscientists needed better tools to manipulate specific nerve cells in the. Two intersections attempting to make a splitgal4 driver line specific for the lplc1 cell type. The trip recommends the mtd gal4 driver for maternal and germline expression of shrnas. In january 2011, the rubin lab began depositing our collection of gal4 driver lines in the bloomington stock center at the rate of approximately 100 lines a week and all lines were available for distribution by december 31, 2012. Our initial screen was designed to examine ubiquitous inhibition of mirna activity using the tub gal4 driver. The gmr gal4 driver, which expresses the yeast transcription factor gal4 under the control of glass multiple reporter gmr promoter elements, has been commonly utilized to express target transgenes, specifically in the developing eye. Development of germ cell specific gal4 driver transgenes has facilitated investigation of gene function in gscs and spermatogonia but specific gal4. To address this need, we assembled a genetic toolkit that allows for an exquisite level of control over the da neural network in drosophila.

Pipeline for generation 1 gal4 driver production and imaging. May 16, 20 building up a library of different gal4 lines using enhancer traps that i will talk about another time allows a wide range of expression patterns to be tested with a single uas controlled gene that only has to be created in the lab a single time. To do this, drosophila geneticists use the gal4uas system. Friedrich,1 igor siwanowicz,1 and hiromu tanimoto1,2 1max. A genetic toolkit for dissecting dopamine circuit function in. Facilitating neuronspecific genetic manipulations in drosophila. Gal4driver lines also can be engineered to include the ubiquitous expression of a temperaturesensitive gal80 gal80 ts that reversibly suppresses gal4 activity at 18c and permits gal4 binding to uas at 29c, thereby permitting temporal control. Expression of gal4 in each line is controlled by a different, defined fragment of genomic dna that serves as a transcriptional enhancer. Facilitating neuronspecific genetic manipulations in. A genetic screen supports a broad role for the drosophila.

Gal4 drivers specific for type ib and type is motor neurons in. The gal4uas binary expression control system has been an extremely useful tool in drosophila genetics. Here, we have employed the drosophila genetics to screen for potential interaction partners of drosophila manf dmmanf in vivo. The gal4 uas binary system has been widely used in drosophila melanogaster for ectopic expression of transgenes in a tissuespecific manner. The flylight project has generated around 7,000 transgenic drosophila lines, in each of which expression of gal4 is controlled by a different transcriptional enhancer that often drives. Theoretically, any gal4 driver can be adapted to generate a lexa or qf driver by using the transposon replacement technique or by borrowing the key celltype specific gene regulatory fragments. The driver lines provide tissuespecific gal4 expression and the responder. A gal4driver line resource for drosophila neurobiology ncbi. Driver lines expressing the transcriptional activator geneswitch in a. Here we describe gal4 drivers specific for examples of neurons within each type, ib or is. Confocal images of the central nervous system from female and male flies carrying the indicated split gal4 driver lines as well as uasmyrflag or uascschrimsonmvenus. Apr 22, 2016 this video describes the fundamental concepts behind the gal4 uas system in drosophila.

New tools for the analysis of glial cell biology in drosophila. The gal4 expression information is provided by the donor of the stock. In these lines referred to below as generation 1 gal4 lines, expression of the transcription factor gal4 is driven by a 23kb segment of genomic dna that contains one or more transcriptional enhancer sequences. Our vienna tiles vt enhancer gal4 lines are typically used in drosophila experiments as drivers to restrict the. Sep 14, 2007 tissue specificity is provided by the choice of gal4 driver.

Jul 20, 2018 border cell bc migration during drosophila oogenesis is an excellent model for the analysis of the migratory and invasive cell behavior. The drosophila testis has proven to be a valuable model organ for investigation of germline stem cell gsc maintenance and differentiation as well as elucidation of the genetic programs that regulate differentiation of daughter spermatogonia. The split gal4 lines can be requested from the janelia fly facility by performing a search and adding the desired lines to your cart. Ac expression patterns of the generation 1 gal4 lines r37h04 a, r64g09 b, and r28g07 c observed with insertion of the transgene in attp2 jenett et al. In drosophila, the two components are carried in separate lines allowing for numerous combinatorial possibilities.

Males from the gal4 driver line were crossed to virgin females of the vdrc uas line for all crosses. Crossing these flies with different gal4 driver lines. To date, thousands of transgenic lines have been generated that exhibit distinct gal4 expression patterns during development and are available for analysis2, 3. Geneticists have created genetic varieties of model organisms typically fruit flies, called gal4 lines, each of. Expression pattern of sev gal4 driver parallels that of. For the fruit fly drosophila melanogaster, one of the most versatile methods to. Nov 01, 2015 fruit flies of the genus drosophila have been an attractive and effective genetic model organism since thomas hunt morgan and colleagues made seminal discoveries with them a century ago. To date, thousands of transgenic lines have been generated that exhibit distinct gal4. Reverse genetics in drosophila dominant negative gal4 uas based rnai injection, gal4 uas based homologous recombinationtilling. Pdf a gal4driver line resource for drosophila neurobiology. Rnai lines were obtained from the vienna drosophila resource center and the gal4 driver lines from the bloomington, indiana drosophila stock. A drosophila model of fusrelated neurodegeneration reveals genetic interaction between fus and tdp43. Gal4 drivers specific for type ib and type is motor neurons. Jan 01, 2020 before initiating a genetic screen of mirsp strains, we compared baseline locomotion in the adult assay with standard wild type control strains cantons and w 1118 and the gal4 driver strains to assess the effects of genetic background.

To facilitate largescale functional studies in drosophila, the drosophila transgenic rnai project trip at harvard medical school hms was established along with several goals. This elegant and robust approach relies on the availability of socalled gal4 driver lines. To limit the number of results displayed in the table below, you can navigate to gal4 regulated by. A promising approach for this is the geneswitch gal4 system of drosophila. Gal4uas targeted gene expression for studying drosophila. The driver and target uas fly lines are crossed, and the full complement of genetic. Expression of gal4 in each line is controlled by a different.

A drosophila model of fusrelated neurodegeneration. All vdrc uas lines were crossed with the fullbody tubulin gal4 sb driver and a pannier gal4 driver y 1 w 1118. Interspecific functional complementation test in drosophila. Thfc tsinghua university beijing, china vdrc, vienna drosophila resource center. Gal4 driverx epuasinsertion lines uas endogenous gene. Genetic reagents for making splitgal4 lines in drosophila. Importantly this allowed systems to be divided so a fly line with gal4 protein being produced with a specific expression pattern can be crossed to a line. These ep or gs elements need to be activated by a gal4 driver line and often result in dominant phenotypes that can be assessed in the first generation of offspring flies. A genetic toolkit for dissecting dopamine circuit function. Lines expressing the gal4 protein are crossed to lines carrying a target gene of interest mbnl1 kiaa0428 in this rescue experiment, subcloned downstream from five gal4 binding sites the uas. In instances of lethality with the tubulin gal4 sb driver, the lines were crossed to another fullbody ubiquitin.

The commonly used eyespecific sev gal4 and gmr gal4 drivers in drosophila melanogaster are expressed in tissues other than eyes also. The pipeline we are using to produce the lines is shown in the first diagram and an example of the analysis as applied to the octopamine receptor 2 gene is shown second. Genetic dissection of active forgetting in labile and. Gal4uas targeted gene expression for studying drosophila hedgehog signaling. The dgrp lines are maintained in the mackay laboratory. The mushroom body of adult drosophila characterized by. Gal4 enhancer traps that can be used to drive gene. Spatial and temporal control of gene expression in drosophila. Another boon to the system of drosophila is the tool for over expression gain of function studiesfor targeted gene expression.

We observed extensive colocalization of our pale fish probes with the r58e02 and ss02425 driver lines figure 2, c and d, and file s9. Genetic reagents for making split gal4 lines in drosophila. Forward genetics in drosophila any mutagenzygotic screen e. Eyes from the line expressing fus wt showed almost normal morphology by light microscopy and histology. Bid transgene, generation of transgenic xy lines, and construction of the ey gal4 cyo. We report here the generation of over 18,000 driver lines the vt collection that exploit the gal4, lexa, and split gal4 systems to express transgenes in distinct and highly specific cell types in drosophila. A genomewide transgenic rnai library for conditional gene. The yeast transcriptional activator gal4 has been extensively utilized as a genetic reporter in drosophila1. Rubin and colleagues have established a collection of 7,000 gal4 driver lines in drosophila. The first is necessary for random insertion in the genome, the latter is necessary for identification of the spatial regulation by the enhancer. Genetics of cocaine and methamphetamine consumption and. Jul 12, 2007 vast collections of gal4 driver lines are already available to target rnai to specific cells 23, and inducible gal4 systems can be used to further restrict rnai to a selected stage of the flys. We established a collection of 7,000 transgenic lines of drosophila melanogaster.

Vp16 made by pernille rorth rorth, 1998 pcog vector robinson and cooley, 1997 contains a promoter from the otu gene and the 3 utr from the k10 gene. The construct is introduced into the drosophila genome with an artificial pelement genetically marked with a drosophila white minigene. Work with drosophila has enabled dramatic advances in cell and developmental biology, neurobiology and behavior, molecular biology, evolutionary and population genetics, and other fields. Border cell bc migration during drosophila oogenesis is an excellent model for the analysis of the migratory and invasive cell behavior. In vivo screening reveals interactions between drosophila.

A the e22c gal4 line drives gfp expression in the follicular stem cells and. Micrornas regulate multiple aspects of locomotor behavior in. We have generated and characterized a series of gal4, gal80, split gal4, killer zipper, homologyassisted crispr knockin hack, flippase flp, and microrna transgenic lines for this purpose. A conditional tissuespecific transgene expression system using. The driver lines provide tissuespecific gal4 expression and the responder lines carry the coding sequence for the gene of interest under the control of uas sites. An illustration of the gal4 uas system from wimmer 2003. Genetic odyssey to generate marked clones in drosophila. Hibbard, view orcid profile amanda cavallaro, juichun kao and view orcid profile gerald m. To generate genetic tools to identify and manipulate motor neurons of each type, we screened for gal4 driver lines for this purpose. Gal80 ts reversibly suppresses gal4 activity at 18c and permits gal4 binding to uas at 29c.

The drosophila melanogaster larval neuromuscular system is extensively used by. Ac expression patterns of the generation 1 gal4 lines r37h04 a. In vivo screening reveals interactions between drosophila manf and genes involved in the mitochondria and the ubiquinone synthesis pathway. When females carrying a uas responder uasgfp are mated to males carrying a gal4 driver progeny containing both elements of the system are produced. The gal4uas system is a biochemical method used to study gene expression and function in. Lgal4 is a strong neuronal cellspecific driver 51, 52, 53, routinely used for genefunction analysis studies in drosophila nervous system.

The trip recommends the mtdgal4 driver for maternal and germline expression of shrnas. Gal4 drivers specific for type ib and type is motor. We document the expression patterns of over 14,000 of these lines in the adult male brain. Neural circuitry linking mating and egg laying in drosophila. When gerry rubin became janelias executive director, he began looking for a research project that fit the campuss focus on imaging and neuroscience. My understanding is that gal4 recruits rna polymerase ii when it binds to the uas with its dnabinding bottom domain, leaving its activation domain top open. Identification of novel negative cellcycle regulators in drosophila through a gainoffunction screen. Using uas gal4 system one can over express a gene of interest in tissue of choice. We used confocal microscopy of dissected nervous systems to determine the expression patterns driven by each fragment in the.

On top of this, the construct usually includes a genetic marker, e. The enhancer trap construct contains a transposable element and a reporter gene. Gal4 is expressed not only in the iscs but also in the eb cells micchelli and perrimon, 2006. It also describes how this genetic tool can be used to knock down or overexpress particular protein in a. Two intersections attempting to make a split gal4 driver line. To further refine targeting of specific da neurons, we also created. A gal4driver line resource for drosophila neurobiology. Genetic tools for manipulating transgene expression. Gal4 is the only gal4 line widely used to manipulate gene expression in the iscs and label the isc. The yeast transcriptional activator gal4 has been extensively used as a genetic reporter in drosophila 1. New slbo gal4 driver lines for the analysis of border cell. Fly light splitgal4 driver collection janelia research campus. This report describes a set of over 2,800 transgenic lines for use with the split gal4 intersectional method. Xue shanghai key laboratory for signaling and diseases, school of life science and technology, tongji university, shanghai, china.

Fus r518k and fus r521c were expressed by using the gmr gal4 driver. The advantage of the gal80 ts system is that it can be combined with existing gal4 drivers and uas effectors, whereas the geneswitch gal4 and gal4 er systems require the creation of new tissuespecific geneswitch gal4 or gal4 er driver lines. Uas, together with a specific gene of interest, is kept in one fly line, and gal4. The gal4 driver lines were tested for the expression of uasgfp. View orcid profile heather dionne, view orcid profile karen l. Trangenic drosophila rnai libraries, a collection of enhancer gal4 driver lines, the tagged flyfos transgeneome ftrg library. Facilitating neuronspecific genetic manipulations in drosophila melanogaster using a split gal4 repressor. This video describes the fundamental concepts behind the gal4uas system in drosophila. This video describes the fundamental concepts behind the gal4 uas system in drosophila. To identify novel regulators of tissue growth, each ep line was crossed to flies expressing ey gal4 and the subsequent progeny were screened for reduced eye size. In turn, gal4 should only activate gene transcription where a uas has been introduced. For study in drosophila, the gal4 gene is placed under the control of a native gene promoter, or driver gene, while the uas controls expression of a target gene.

For study in drosophila, the gal4 gene is placed under the control of a native gene promoter, or driver gene, while. A the e22c gal4 line drives gfp expression in the follicular stem cells and thus in all fcs, albeit patchy. Reverse genetics in drosophila dominant negative gal4 uas based. All lines were maintained on standard cornmealyeastmolasses medium at 25c on a 12 hour lightdark cycle with constant humidity of 50%. New slbo gal4 driver lines for the analysis of border. All vdrc uas lines were crossed with the fullbody tubulin gal4 sb driver and a pannier gal4 driver. To address this issue, we present here a set of genetic tools designed to facilitate the functional dissection of da neural networks in drosophila. Genetic architecture of abdominal pigmentation in drosophila. Rnai lines were obtained from the vienna drosophila resource center and the gal4 driver lines from the bloomington, indiana drosophila stock center.

National institute of genetics mishima, japan tsinghua fly center thfc transgenic rnai fly lines. Pdf genetic reagents for making splitgal4 lines in. A different, defined fragment of genomic dna serves as a transcriptional enhancer to control gal4 expression in each line. Pdf the commonly used eyespecific sevgal4 and gmrgal4.