A NON-RADIOACTIVE INSITU HYBRIDIZATION METHOD FOR THE LOCALIZATION OF SPECIFIC RNAS IN DROSOPHILA EMBRYOS REVEALS TRANSLATIONAL CONTROL OF THE SEGMENTATION GENE HUNCHBACK
A NON-RADIOACTIVE INSITU HYBRIDIZATION METHOD FOR THE LOCALIZATION OF SPECIFIC RNAS IN DROSOPHILA EMBRYOS REVEALS TRANSLATIONAL CONTROL OF THE SEGMENTATION GENE HUNCHBACK
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DOI：
10.1007/bf00291041
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发表时间：
1989-08-01
期刊：
影响因子：
1.6
通讯作者：
PFEIFLE, C
中科院分区：
文献类型：
--
作者：
TAUTZ, D;PFEIFLE, C
我们开发了一种非放射性原位杂交技术，用于在整装果蝇胚胎中对RNA进行定位。固定后，整个胚胎与用地高辛标记的DNA探针进行原位杂交。通过使用一种抗地高辛的磷酸酶偶联抗体来检测杂交产物。在平行实验中，可以用一种针对相应蛋白质产物的抗体处理胚胎，以便利用标准的免疫化学技术检测其分布。我们利用这种方法比较了在胚胎发生早期阶段分节基因驼背（hb）的RNA和蛋白质产物的时空分布模式。这种比较揭示了母源hb mRNA的翻译调控，这用常规技术是难以检测到的。非放射性原位杂交方法与常规方法一样灵敏，但速度更快且更易于操作。这可能使其成为适用于多种其他系统的有用工具。
We have developed a non-radioactive in situ hybridization technique for the localization of RNA in whole mount Drosophila embryos. After fixation, whole embryos are hybridized in situ with a DNA probe which has been labeled with digoxygenin. The hybridization products are detected by using a phosphatase-coupled antibody against digoxygenin. In parallel experiments, embryos can be treated with an antibody directed against the corresponding protein product to allow the detection of its distribution using standard immunochemical techniques. We have used this approach to compare the spatial and temporal distribution patterns of the RNA and protein products of the segmentation gene hunchback (hb) during the early stages of embryogenesis. This comparison revealed translational control of the maternally derived hb mRNA, which was difficult to detect by conventional techniques. The non-radioactive in situ hybridization method is as sensitive as conventional methods, but is faster and easier to perform. This may make it a useful tool for a variety of other systems.