Consistent transient transfection of DNA into non-transformed human and murine T-lymphocytes

Academic Article

Abstract

  • The ability to analyze transcriptional regulation in non-transformed T- cells has been hampered by the inability to reproducibly transiently transfect these cells with DNA constructs. We have previously demonstrated that normal human whole mononuclear and CD4 T-cells can be consistently transiently transfected with plasmid DNA. Human cells were most receptive to plasmid DNA uptake between 19.5 and 20 h after prestimulation with a submitogenic dose of the polyclonal T-cell activator, PHA. Here, we report an alteration and optimization of this protocol for non-transformed murine splenic T-cells, using concanavalin A instead of PHA as the preactivation stimulus. When coupled with the high sensitivity of luciferase reporter gene constructs, this protocol facilitates the analysis of a variety of T-cell- specific promoters in non-transformed T-cells. In addition, we directly demonstrate that murine T-cells are specifically transiently transfected among a population of whole mononuclear cells by using an expression vector for green fluorescent protein.
  • Published In

    Digital Object Identifier (doi)

    Author List

  • Cron RQ; Schubert LA; Lewis DB; Hughes CCW
  • Start Page

  • 145
  • End Page

  • 150
  • Volume

  • 205
  • Issue

  • 2