Resources for functional studies in Drosophila

项目总结/摘要 从基于scRNAseq的研究中获得的最新知识为研究提供了新的机会。 表征细胞类型和描述表型。此外，从scRNAseq获得的信息 可以帮助开发用于功能研究的新试剂。scRNAseq研究的主要挑战之一是 要解决的是scRNAseq簇到解剖结构的映射。虽然这可以相对容易地完成，如果一个集群 由单个差异表达基因（DEG）的独特表达定义，任务更多 如果一组单元格只能使用DEG的组合来定义，则会很复杂。要识别以下组合， 基因的表达定义细胞簇，我们在目标1中提出开发一种算法， 定义一个簇的最小唯一标记基因集。一些方法和资源可以 用于将标记基因的表达与解剖结构相关联;然而，在大多数情况下， 需要多个基因来将簇正确地映射到解剖结构，并且现有的试剂是有限的。为 分裂Gal 4方法，其中Gal 4的DNA结合结构域和转录激活因子结构域被 在两种不同增强子的控制下独立表达，可扩展用于检测多种 基因，在目标2中，我们建议建立在我们建立的TRiP/DRSC平台上， 的1012个分裂-Gal 4系，其将用于映射存在于果蝇中的估计的450个细胞簇。 重要的是，从这些研究中产生的分裂Gal 4系将使我们能够解决一个长期存在的问题， 果蝇领域，即现有Gal 4系的特异性，因为我们和其他人已经观察到， 大多数所谓“组织特异性”Gal 4驱动细胞系也在另外的不需要的细胞中表达 类型或组织。最后，由于目前的分裂Gal 4系统与时间控制不兼容， 使用分裂Gal 4系进行需要UAS驱动的转基因的阶段特异性活化的实验， 目标3，我们将开发一个强大的分裂Gal 4方法，允许时间控制，并产生一个 200个可诱导的分裂Gal 4系的集合，其将允许产生100个可控组织 特定的Gal 4模式，覆盖大多数幼虫和成虫组织。这些资源将共同提供 社区急需的试剂，将尽快提供和分发， 由布卢明顿果蝇库存中心生产。

PROJECT SUMMARY / ABSTRACT Recent knowledge gained from studies based on scRNAseq provides new opportunities for the characterization of cell types and description of phenotypes. In addition, information gained from scRNAseq can help develop new reagents useful for functional studies. One of the main challenges in scRNAseq studies to address is the mapping of scRNAseq clusters to anatomy. While this can be relatively easily done if a cluster is defined by the unique expression of a single differentially expressed gene (DEG), the task is more complicated if a group of cells can only be defined using a combination of DEGs. To identify combinations of genes whose expression define cell clusters, we propose in Aim 1 to develop an algorithm that determines the smallest unique set of marker genes that define a cluster. A number of approaches and resources can be used to associate the expression of marker genes to the anatomy; however, in most cases co-expression of multiple genes is required to properly map clusters to the anatomy, and existing reagents are limited. As the split-Gal4 method, whereby the DNA-binding domain of Gal4 and a transcriptional activator domain are expressed independently under the control of two different enhancers, is scalable for the detection of multiple genes, in Aim 2, we propose to build on our established TRiP/DRSC platform and generate a collection of 1012 split-Gal4 lines that will be used to map the estimated 450 cell clusters present in the fly. Importantly, the split-Gal4 lines generated from these studies will allow us to address a long-standing issue in the Drosophila field, namely the specificity of existing Gal4 lines, as we and others have observed that the large majority of so-called “tissue-specific” Gal4 driver lines are also expressed in additional, unwanted cell types or tissues. Finally, as the current split-Gal4 system is not compatible with temporal control, preventing the use of split-Gal4 lines for experiments requiring stage specific activation of the UAS driven transgene, in Aim 3, we will develop a robust split-Gal4 method that allows temporal control and generate a collection of 200 inducible split-Gal4 lines that will allow the generation of 100 controllable tissue specific Gal4 patterns, covering most larval and adult tissues. Together, these resources will provide the community with much needed reagents that will be made available and distributed as soon as they are produced by the Bloomington Drosophila Stock Center.