Godwin A R, Stadler H S, Nakamura K, Capecchi M R
Howard Hughes Medical Institute, Department of Human Genetics, University of Utah School of Medicine, Salt Lake City, UT 84112, USA.
Proc Natl Acad Sci U S A. 1998 Oct 27;95(22):13042-7. doi: 10.1073/pnas.95.22.13042.
The ability to use a vital cell marker to study mouse embryogenesis will open new avenues of experimental research. Recently, the use of transgenic mice, containing multiple copies of the jellyfish gene encoding the green fluorescent protein (GFP), has begun to realize this potential. Here, we show that the fluorescent signals produced by single-copy, targeted GFP in-frame fusions with two different murine Hox genes, Hoxa1 and Hoxc13, are readily detectable by using confocal microscopy. Since Hoxa1 is expressed early and Hoxc13 is expressed late in mouse embryogenesis, this study shows that single-copy GFP gene fusions can be used through most of mouse embryogenesis. Previously, targeted lacZ gene fusions have been very useful for analyzing mouse mutants. Use of GFP gene fusions extends the benefits of targeted lacZ gene fusions by providing the additional utility of a vital marker. Our analysis of the Hoxc13(GFPneo) embryos reveals GFP expression in each of the sites expected from analysis of Hoxc13(lacZneo) embryos. Similarly, Hoxa1(GFPneo) expression was detected in all of the sites predicted from RNA in situ analysis. GFP expression in the foregut pocket of Hoxa1(GFPneo) embryos suggests a role for Hoxa1 in foregut-mediated differentiation of the cardiogenic mesoderm.
利用一种重要的细胞标记物来研究小鼠胚胎发生的能力将开辟实验研究的新途径。最近，含有编码绿色荧光蛋白（GFP）的水母基因多个拷贝的转基因小鼠的应用已开始实现这一潜力。在此，我们表明，通过共聚焦显微镜很容易检测到单拷贝、靶向的GFP与两个不同的小鼠Hox基因Hoxa1和Hoxc13的读码框内融合产生的荧光信号。由于Hoxa1在小鼠胚胎发生早期表达，而Hoxc13在晚期表达，本研究表明单拷贝GFP基因融合可用于小鼠胚胎发生的大部分过程。以前，靶向lacZ基因融合对于分析小鼠突变体非常有用。GFP基因融合的应用通过提供重要标记物的额外用途扩展了靶向lacZ基因融合的优势。我们对Hoxc13（GFPneo）胚胎的分析揭示了在Hoxc13（lacZneo）胚胎分析所预期的每个位点中都有GFP表达。同样，在RNA原位分析预测的所有位点中都检测到了Hoxa1（GFPneo）的表达。Hoxa1（GFPneo）胚胎前肠袋中的GFP表达表明Hoxa1在前肠介导的心脏中胚层分化中起作用。
