title: An electroporation protocol for efficient DNA transfection in PC12 cells.
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.
authors.affiliation: Laboratory of RNA Biology and Biotechnology, Centre for Integrative Biology (CIBIO), University of Trento, via delle Regole 101, 38123, Trento, Italy.
authors.forename: Giuseppina
authors.initials: G
authors.lastname: Covello
authors.forename: Kavitha
authors.initials: K
authors.lastname: Siva
authors.forename: Valentina
authors.initials: V
authors.lastname: Adami
authors.forename: Michela A
authors.initials: MA
authors.lastname: Denti