水稻RNA原位杂交体系的优化

为了能准确、快捷地追踪目标基因在水稻中的表达,并提供在不同植物中基因表达的研究基础,优化了水稻RNA原位杂交体系。以卡诺固定液处理材料,原位杂交结果显示:细胞边界清楚,标本背景清晰,检测信号强。设置了0、15、25、35和45 min 5个消化时间梯度,消化时间是25 min的试验结果显示:杂交信号分布于整个受检表面,信号可检测性强,并在不同组织结构中信号强弱不同。以根尖为材料,杂交液A的检测信号符合分布规律,杂交液B验证反义探针检测信号较弱,杂交液C验证反义探针检测信号进一步减弱,并伴随着验证正义探针信号增强。以叶片为试验材料,杂交液A和B的原位杂交结果差异不大,而杂交液C验证反义探针信号减弱,验证正义探针信号增强。结论:水稻RNA原位杂交体系的固定液为卡诺固定液,最佳消化时间为25 min,杂交液为杂交液A。

Optimization of the RNA in situ hybridization system in rice

In order to track the target gene expression in rice accurately and quickly,and to provide research base for gene expression in different plants.The paper optimized RNA in situ hybridization system.The result of Canony fixative showed that the cell borders were seen clearly,background was clean,and the testing signal was recognizable.Canony fixative is preferred to the others for experiments.The experimental set digestion gradient time:0,15,25,35 and 45 min.The results of 25 min showed that hybridization signal was distributed throughout the surface of paraffin slice,and a strong signal could be detected.In different organizational structures,the signal displayed different blue colors.The result indicated that in the different organizations,the number of expressed RNA is different.This study determined 25 min for the optimal digestion time.In the root tip,hybrid solution A showed that the signal distribution was consistent with regular pattern,hybrid solution B showed that the antisense signal was weaker than A,and hybridization solution C had a further weaker antisense signal and deeper blue sense signal.In rice leaves,hybridization solution A and B had similar results,the hybridization solution C weakened the antisense signal while the sense signal was enhanced.So hybridization solution A was determined as the preferred hybridization solution.