Department of Agrobiology and Bioresources, Faculty of Agriculture, Utsunomiya University, Utsunomiya, Tochigi, Japan.
Institute of Agrobiological Sciences, National Agriculture and Food Research Organization, Tsukuba, Ibaraki, Japan.
Methods Mol Biol. 2025;2869:7-13. doi: 10.1007/978-1-0716-4204-7_2.
Ribonucleic acid (RNA) extraction is the first critical step in gene expression analysis. In this chapter, we describe a high-throughput RNA extraction method using guanidine thiocyanate and isopropyl alcohol (HighGI). The use of carboxyl-coated paramagnetic beads, instead of silica membrane columns, enables semi-automation using a liquid handling system and high-throughput RNA extraction for large-scale transcriptome studies. Homemade mixes of paramagnetic beads and buffers make HighGI inexpensive. In addition, HighGI-extracted RNA retains low molecular weight RNA molecules less than 200 bp, which is typically lost in commercial column-based kits.
RNA 提取是基因表达分析的关键步骤之一。在本章中，我们描述了一种使用异硫氰酸胍和异丙醇（HighGI）的高通量 RNA 提取方法。使用羧基涂层的超顺磁珠代替硅胶膜柱，可使用液体处理系统进行半自动化，并可进行大规模转录组研究的高通量 RNA 提取。超顺磁珠和缓冲液的自制混合物使 HighGI 价格低廉。此外，HighGI 提取的 RNA 保留了低分子量 RNA 分子，小于 200 bp，这通常在商业柱基试剂盒中丢失。
