Cite as: Cold Spring Harb. Protoc.; 2006; doi:10.1101/pdb.prot3871

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Calcium-phosphate-mediated Transfection of Eukaryotic Cells with Plasmid DNAs

Joseph Sambrook and David W. Russell

This protocol was adapted from Molecular Cloning, 3rd edition, by Joseph Sambrook and David W. Russell. Cold Spring Harbor Laboratory Press, Cold Spring Harbor, NY, USA, 2001

The first 100 words of the full text of this article appear below.


INTRODUCTION

Calcium phosphate forms an insoluble precipitate with DNA, which attaches to the cell surface and is taken into the cells by endocytosis. This protocol is a modified version of a method published by Jordan et al. (1996) who rigorously optimized calcium-phosphate-based transfection methods for Chinese hamster ovary cells and the 293 line of human embryonic kidney cells. The protocol is easily adapted for use with other types of cells, both adherent and nonadherent.


MATERIALS

recipe 0.1x TE (pH 7.6)

recipe 2x HEPES-buffered saline

recipe caution CaCl2 (2.5 M)

Cell culture growth medium (complete and [optional] selective)

recipe Chloroquine (100 mM) (optional)

Dissolve 52 mg of chloroquine . . . [Full Text of this Article]


METHOD


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