Voutev Roumen, Hubbard E Jane Albert
Department of Biology, Developmental Genetics Program, New York University, New York, New York 10003, USA.
Genetics. 2008 Sep;180(1):103-19. doi: 10.1534/genetics.108.090274. Epub 2008 Aug 24.
We present a two-part system for conditional FLP-out of FRT-flanked sequences in Caenorhabditis elegans to control gene activity in a spatially and/or temporally regulated manner. Using reporters, we assess the system for efficacy and demonstrate its use as a cell lineage marking tool. In addition, we construct and test a dominant-negative form of hlh-12, a gene that encodes a basic helix-loop-helix (bHLH) transcription factor required for proper distal tip cell (DTC) migration. We show that this allele can be conditionally expressed from a heat-inducible FLP recombinase and can interfere with DTC migration. Using the same DTC assay, we conditionally express an hlh-12 RNAi-hairpin and induce the DTC migration defect. Finally, we introduce a set of traditional and Gateway-compatible vectors to facilitate construction of plasmids for this technology using any promoter, reporter, and gene/hairpin of interest.
我们提出了一种两部分系统,用于在秀丽隐杆线虫中以空间和/或时间调控的方式条件性地敲除FRT侧翼序列,以控制基因活性。我们使用报告基因评估该系统的有效性,并证明其作为细胞谱系标记工具的用途。此外,我们构建并测试了hlh-12的显性负性形式,hlh-12是一个编码适当的远端末梢细胞(DTC)迁移所需的基本螺旋-环-螺旋(bHLH)转录因子的基因。我们表明,该等位基因可以从热诱导的FLP重组酶中条件性表达,并能干扰DTC迁移。使用相同的DTC检测方法,我们条件性表达hlh-12 RNAi发夹并诱导DTC迁移缺陷。最后,我们引入了一组传统的和与Gateway兼容的载体,以促进使用任何感兴趣的启动子、报告基因和基因/发夹构建用于该技术的质粒。