BioTechniques (Jan 2002)

Efficacy of an Amphipathic Oligopeptide to Shuttle and Release a cis-Acting DNA Decoy into Human Cells

  • L. Citti,
  • P. Rovero,
  • M.G. Colombo,
  • L. Mariani,
  • L. Poliseno,
  • G. Rainaldi

DOI
https://doi.org/10.2144/02321dd03
Journal volume & issue
Vol. 32, no. 1
pp. 172 – 177

Abstract

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We investigated the ability of an amphipathic oligopeptide to carry a synthetic ds- DNA oligonucleotide inside human cells. The oligonucleotide was designed as a decoy binding site for the transcriptional activator of the methylguanine-DNA methyltransferase (MGMT) gene. The complex oligopeptide and decoy were administered to MCF10A exponentially growing cells, and the uptake was monitored by flow cytometry. After a 1-h exposure, almost all of the MCF10A cells were fluorescent, indicating that all of the cells had been transfected. By increasing the time, the fluorescence intensity per cell rapidly increased to a plateau at the 8-h time point. RT-PCR analysis of the MGMT gene was used as the molecular readout of the intracellular activity of the DNA decoy. MCF10A cells transfected with the oligopeptide/decoy complex showed a strong reduction in MGMT mRNA. Here, we discuss the advantages of using amphipathic oligopeptides as carriers of short DNA sequences.