Hernández-Castellano Sara, Nic-Can Geovanny I, De-la-Peña Clelia
Unidad de Biotecnología, Centro de Investigación Científica de Yucatán, Calle 43 No. 130, Col. Chuburná de Hidalgo, Merida, Yucatán, 97200, Mexico.
Campus de Ciencias Exactas e Ingeniería, Universidad Autónoma de Yucatán, Periférico Norte. Km 33.5, Tablaje catastral 13615 Col. Chuburná de Hidalgo In, Merida, Yucatán, CP 97200, Mexico.
Methods Mol Biol. 2017;1456:51-62. doi: 10.1007/978-1-4899-7708-3_4.
Among the epigenetic mechanisms studied with a greater interest in the last decade are the microRNAs (miRNAs). These small noncoding RNA sequences that are approximately 17-22 nucleotides in length play an essential role in many biological processes of various organisms, including plants. The analysis of spatiotemporal expression of miRNAs provides a better understanding of the role of these small molecules in plant development, cell differentiation, and other processes; but such analysis is also an important method for the validation of biological functions. In this work, we describe the optimization of an efficient protocol for the spatiotemporal analysis of miRNA by in situ hybridization using different plant tissues embedded in paraffin. Instead of LNA-modified probes that are typically used for this work, we use conventional oligonucleotide probes that yield a high specificity and clean distribution of miRNAs.
在过去十年中受到更多关注的表观遗传机制中，微小RNA（miRNA）是其中之一。这些长度约为17 - 22个核苷酸的小型非编码RNA序列在包括植物在内的各种生物体的许多生物学过程中发挥着重要作用。对miRNA的时空表达分析有助于更好地理解这些小分子在植物发育、细胞分化和其他过程中的作用；而且这种分析也是验证生物学功能的重要方法。在这项工作中，我们描述了一种高效方案的优化，该方案用于通过原位杂交对石蜡包埋的不同植物组织进行miRNA的时空分析。我们使用常规寡核苷酸探针，而非通常用于此项工作的锁核酸（LNA）修饰探针，这些常规探针能产生高特异性且miRNA分布清晰的结果。
