Drea Sinéad, Derbyshire Paul, Koumproglou Rachil, Dolan Liam, Doonan John H, Shaw Peter
Department of Molecular, Cell and Developmental Biology, Yale University, New Haven, CT 06520, USA.
Methods Mol Biol. 2009;513:229-42. doi: 10.1007/978-1-59745-427-8_12.
In the post-genomic era, it is necessary to adapt methods for gene expression and functional analyses to more high-throughput levels of processing. mRNA in situ hybridization (ISH) remains a powerful tool for obtaining information regarding a gene's temporal and spatial expression pattern and can therefore be used as a starting point to define the function of a gene or a whole set of genes. We have deconstructed 'traditional' ISH techniques described for a range of organisms and developed protocols for ISH that adapt and integrate a degree of automation to standardized and shortened protocols. We have adapted this technique as a high-throughput means of gene expression analysis on wax-embedded plant tissues and also on whole-mount tissues. We have used wax-embedded wheat grains and Arabidopsis floral meristems and whole-mount Arabidopsis roots as test systems and show that it is capable of highly parallel processing.
在后基因组时代，有必要使基因表达和功能分析方法适应更高通量的处理水平。mRNA原位杂交（ISH）仍然是获取有关基因时空表达模式信息的有力工具，因此可作为确定单个基因或一组基因功能的起点。我们解构了针对一系列生物体描述的“传统”ISH技术，并开发了ISH方案，该方案将一定程度的自动化应用于标准化和缩短的方案中。我们将该技术改编为对蜡包埋植物组织以及整装组织进行基因表达分析的高通量方法。我们使用蜡包埋的小麦籽粒、拟南芥花分生组织和整装拟南芥根作为测试系统，并表明它能够进行高度并行的处理。
