RNAi expression vectors in mammalian cells

Methods Mol Biol. 2004:252:483-91. doi: 10.1385/1-59259-746-7:483.

Abstract

RNA interference (RNAi) is a recently developed technique for gene silencing by introducing dsRNA into cells, and it is shown to work in mammalian cells when siRNAs are used. Several groups have developed vector-based siRNA expression systems that can induce RNAi in living cells. These vector systems use a pol III promoter, such as U6 or H1, and are classified into two groups based on the form of expressed RNAs: tandem-type and hairpin-type. Here, we describe how to generate these siRNA expression vectors and outline the experimental procedure for suppressing the expression of a reporter gene by transient transfection of a siRNA expression vector.

MeSH terms

  • Animals
  • Base Sequence
  • Cell Line
  • DNA Primers
  • Escherichia coli / genetics
  • Genetic Vectors*
  • HeLa Cells
  • Humans
  • Mammals
  • Molecular Sequence Data
  • Nucleic Acid Hybridization
  • Promoter Regions, Genetic / genetics
  • RNA Interference / physiology*
  • RNA, Catalytic / genetics
  • RNA, Small Interfering / genetics*
  • Restriction Mapping
  • Transcription, Genetic

Substances

  • DNA Primers
  • RNA, Catalytic
  • RNA, Small Interfering
  • hairpin ribozyme