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An electroporation protocol for efficient DNA transfection in PC12 cells.

Authors
  • Covello, Giuseppina
  • Siva, Kavitha
  • Adami, Valentina
  • Denti, Michela A
Type
Published Article
Journal
Cytotechnology
Publication Date
Aug 01, 2014
Volume
66
Issue
4
Pages
543–553
Identifiers
DOI: 10.1007/s10616-013-9608-9
PMID: 23846478
Source
Medline
License
Unknown

Abstract

A wide variety of mammalian cell types is used in gene transfection studies. Establishing transfection methods that enable highly efficient DNA uptake has become increasingly important. PC12 is an established rat pheochromocytoma cell line, which responds to exposure to NGF with cessation of growth, expression of cytoplasmic processes, and differentiation into cells resembling sympathetic neurons. Although PC12 cells represent an important model system to study a variety of neuronal functions, they proved relatively difficult to transfect. We have compared the efficiency of three different chemical transfection reagents (Lipofectamine 2000, Lipofectamine LTX and TransIT-LT1) and of two electroporation systems (Neon and Gene Pulser Xcell) in transiently transfecting undifferentiated PC12 cells. By comparing efficiencies from replicate experiments we proved electroporation (in particular Neon) to be the method of choice. By optimizing different parameters (voltage, pulse width and number of pulses) we reached high efficiency of transfection (90 %) and viability (99 %). We also demonstrated that, upon electroporation, cells are not altered by the transfection and maintain their ability to differentiate.

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