Simin Karl, Scuderi Anne, Reamey James, Dunn Diane, Weiss Robert, Metherall James E, Letsou Anthea
Department of Human Genetics, University of Utah, Salt Lake City, Utah 84112, USA.
Genome Res. 2002 Jul;12(7):1040-7. doi: 10.1101/gr.84402.
Here we describe a high-throughput screen to isolate transcripts with spatially restricted patterns of expression in early embryos. Our approach utilizes robotic automation for rapid analysis of sequence-selected cDNAs in a whole-mount in situ hybridization assay. We determined the spatial distribution of a random collection of 778 different genes from an embryonic cDNA library and show that a significant fraction of these exhibit patterned profiles of expression. In addition, gene ontology studies revealed groups of gene products exhibiting shared expression patterns, providing new insights into the largely overlooked effector molecules that function in development. As described in this paper, automated hybridization to whole-mount embryos in situ proved to be straightforward and provided us with a very powerful method for the global survey of gene expression in early embryos. From the perspective of biological significance, our finding that many spatially restricted transcripts correspond to loci encoding novel transcripts that have not been previously identified in nearly saturating genetic screens for maternal effect and zygotic lethals is particularly notable.
在此，我们描述了一种高通量筛选方法，用于分离早期胚胎中具有空间限制表达模式的转录本。我们的方法利用机器人自动化技术，在全胚胎原位杂交实验中对序列选择的cDNA进行快速分析。我们确定了来自胚胎cDNA文库的778个不同基因的随机集合的空间分布，并表明其中很大一部分呈现出模式化的表达谱。此外，基因本体研究揭示了呈现共享表达模式的基因产物组，为发育过程中很大程度上被忽视的效应分子提供了新的见解。如本文所述，全胚胎原位自动杂交被证明是直接可行的，并为我们提供了一种非常强大的方法，用于全面调查早期胚胎中的基因表达。从生物学意义的角度来看，我们的发现尤其值得注意，即许多具有空间限制的转录本对应于编码新转录本的基因座，这些转录本在几乎饱和的母体效应和合子致死基因筛选中尚未被先前鉴定。
