用RNA-RNA原位杂交技术研究Robo2在早期鸡胚发育中的表达

The expression pattern of Robo2 in early chick embryos using RNA-RNA in situ hybridization

目的:探讨Robo2(roundabout homolog 2)在早期鸡胚发育中的表达。方法:利用分子生物学手段,构建Robo2/pSPT18重组质粒,并且制备地高辛标记的Robo2 RNA探针,进而通过RNA-RNA原位杂交技术检测Robo2在早期鸡胚发育中的表达。结果:原条发生前Robo2表达较弱,原条发生后主要表达在原条和神经板。体节期Robo2主要表达在脊索、神经管、体节和血岛部位。冰冻切片后观察到Robo2主要表达在外胚层,而在中胚层和内胚层只有部分表达。结论:阐明了Robo2在早期鸡胚发育中的表达,为进一步研究Robo2在正常生理和病理条件下的功能及作用机制奠定基础。

AIM：To study the expression pattern of roundabout homolog 2（Robo2） in the early chick embryos.METHODS： The plasmids of Robo2/pSPT18 were constructed and the digoxigenin（DIG）-labeled sense and anti-sense RNA probes were produced by in vitro transcription according to a standard protocol.Finally,the expression pattern of Robo2 in the early chick embryos was investigated using RNA-RNA in situ hybridization with the synthesized probes.RESULTS： At pre-streak stage,the expression of Robo2 was weak.After primitive streak formed,Robo2 was mainly expressed in the primitive streak and neural plate.At somite stage,strong expression of Robo2 in the notochord,neural tube,somites and blood island of the embryos was observed.After whole mount in situ hybridization,the embryos were embedded,frozen and cut to 20 μm sections.The results showed that the expression of Robo2 was strong in the epiblast,but weak in the mesoderm and endoderm.CONCLUSION： The expression pattern of Robo2 in early chick embryos obtained in this experiment provides an approach to further study the biological functions and mechanisms of Robo2 in physiological and pathological conditions.