通过RNAi使基因特异性沉默
RNA interference (RNAi) is a phenomenon in which the introduction of double-stranded RNA (dsRNA) into a diverse range of organisms and cell types causes degradation of the complementary mRNA (Figure 1; step 1). In the cell, long dsRNAs are cleaved into short 21-25 nucleotide small interfering RNAs, or siRNAs, by a ribonuclease known as Dicer (step 2). The siRNAs subsequently assemble with protein components into an RNA-induced silencing complex (RISC), unwinding in the process (step 3). Activated RISC then binds to complementary transcript by base pairing interactions between the siRNA antisense strand and the mRNA. The bound mRNA is cleaved (step 4) and sequence specific degradation of mRNA (step 5) results in gene silencing (reviewed in 1-3).
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| Figure 1. RNAi Mechanism. |
RNAi has been used as a tool by scientists to understand gene function inCaenorhabditis elegans and Drosophila. In these organisms, RNAi can be induced by introducing long dsRNA complementary to the target mRNA to be degraded. In mammalian cells and organisms, however, introducing dsRNA longer than 30 bp activates a potent antiviral response. To circumvent this, siRNAs are used to induce RNAi in mammalian cells and organisms.
In the last few years, siRNAs have been used in a number of different experimental settings to silence gene expression. In some, chemically synthesized or in vitro transcribed siRNAs have been transfected into cells, injected into mice, or introduced into plants (e.g. by a particle gun). In others, siRNAs have been expressed endogenously from siRNA expression vectors or PCR products in cells or in transgenic animals.
In addition to their role in gene silencing, siRNAs have been determined to play diverse biological functions in vivo -- roles that include antiviral defense, transposon silencing, gene regulation, centromeric silencing, and genomic rearrangements. This functional diversity has exemplified the importance of siRNAs within cells and has also stirred interest in their detection across species and tissues.
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