期刊
CHEMBIOCHEM
卷 4, 期 10, 页码 928-935出版社
WILEY-V C H VERLAG GMBH
DOI: 10.1002/cbic.200300708
关键词
antisense agents; gene expression; interference; knockdown; RNA
The postgenomic era is characterized by an almost intimidating amount of information regarding the sequences and expression of previously unknown genes. In response, researchers have developed an increasing interest in functional studies. At the start of such a study, one may have little more than sequence information and bioinformatic annotation. The next step is to hypothesize a potential role in the contest of a cell. Testing of the hypothesis needs to be fast, cheap, and applicable to a large number of genes. Knockdown methods that rely on binding of antisense oligonucleotides to mRNA combined with a subsequent functional assay in cell culture fulfill these requirements: sequence information is sufficient for synthesis of active inhibitors. Depending on the in vitro model chosen, knockdown of gene expression can be achieved with medium or even high throughput. The two most popular methods of knockdown in cell culture are the use of antisense oligonucleotides that rely on ribonuclease H (RNAse H)-dependent cleavage of mRNA, and RNA interference triggered by small double-stranded RNA molecules. Both methods act in a sequence-specific manner and can give efficient knockdown. IN both cases, researchers struggle with nonspecific off-target effects and the difficulty of site selection. Studies that compare the methods differ in their judgment as to which method is superior.
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