Targeted gene expression without a tissue-specific promoter: Creating mosaic embryos using laser-induced single-cell heat shock
Targeted gene expression without a tissue-specific promoter: Creating mosaic embryos using laser-induced single-cell heat shock
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DOI：
10.1073/pnas.94.12.6255
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发表时间：
1997-06-10
影响因子：
11.1
通讯作者：
Keshishian, H
中科院分区：
文献类型：
--
作者：
Halfon, MS;Kose, H;Keshishian, H
我们已经开发出一种方法，将果蝇胚胎中的基因表达靶向到特定细胞，而无需在该特定细胞中指导表达的启动子。使用数字增强成像系统来识别活胚胎中的单个细胞，我们通过使用激光微束对每个细胞单独进行热休克。1至2分钟的激光处理足以诱导热休克反应，但对热休克细胞不致命。通过hsp 26-lacZ报告构建体的β-半乳糖苷酶的表达或通过hsGAL 4诱导后UAS靶基因的表达，在多种细胞类型（包括神经元和体细胞肌肉）中测量热休克的诱导。我们讨论了这种技术异位基因表达的研究，谱系追踪，基因失活的研究，并在体外细胞的研究的适用性。激光热休克是一种通用技术，可适用于各种研究生物体，并且对于希望仅在不同细胞或细胞克隆中瞬时或组成性表达给定基因的任何研究都是有用的。在选择的时间点。
We have developed a method to target gene expression in the Drosophila embryo to a specific cell without having a promoter that directs expression in that particular cell. Using a digitally enhanced imaging system to identify single cells within the living embryo, we apply a heat shock to each cell individually by using a laser microbeam. A 1- to 2-min laser treatment is sufficient to induce a heat-shock response but is not lethal to the heat-shocked cells, Induction of heat shock was measured in a variety of cell types, including neurons and somatic muscles, by the expression of beta-galactosidase from an hsp26-lacZ reporter construct or by expression of a UAS target gene after induction of hsGAL4. We discuss the applicability of this technique to ectopic gene expression studies, lineage tracing, gene inactivation studies, and studies of cells in vitro. Laser heat shock is a versatile technique that can be adapted for use in a variety of research organisms and is useful for any studies in which it is desirable to express a given gene in only a distinct cell or clone of cells, either transiently or constitutively, at a time point of choice.