Moela Pontsho
Division of Genetics, Department of Biochemistry, Genetics, and Microbiology, Faculty of Natural and Agricultural Sciences, University of Pretoria, Pretoria, South Africa.
Methods Mol Biol. 2025;2938:109-114. doi: 10.1007/978-1-0716-4607-6_12.
The most common technique used in silencing gene expression adopts a naturally occurring cellular process called RNA interference (RNAi), which uses a double-stranded RNA molecule to silence homologous endogenous genes by degrading their cognate mRNA transcripts. The presence of long double-stranded RNA molecules triggers this process, which ensues by first shortening the lengthy transcripts enzymatically followed by their incorporation into the RISC complex thus allowing it to find and degrade complementary mRNA sequences. In an in vitro setting, short interfering oligonucleotides are commercially synthesized and commonly transfected into mammalian cells through lipofection. This current chapter illustrates the silencing of PPARγ gene using RNAi in adherent mammalian cells.
沉默基因表达最常用的技术采用一种自然发生的细胞过程，称为RNA干扰（RNAi），它利用双链RNA分子通过降解同源内源性基因的同源mRNA转录本来使其沉默。长双链RNA分子的存在触发了这一过程，该过程首先通过酶促作用缩短长转录本，然后将其整合到RISC复合物中，从而使其能够找到并降解互补的mRNA序列。在体外环境中，短干扰寡核苷酸通过商业合成，并通常通过脂质转染法转染到哺乳动物细胞中。本章阐述了在贴壁哺乳动物细胞中使用RNAi沉默PPARγ基因的方法。
