Transfection of KB Cells by Polyethylene Glycol-induced Fusion with Erythrocyte Ghosts Containing Adenovirus Type 2 DNA

Open Access
Publisher Website
Google Scholar
Abstract
Cell ghosts were formed by hypotonic haemolysis and subsequent isotonic re-sealing of human erythrocytes in the presence of adenovirus type 2 DNA. The ghosts entrapped virus DNA with an efficiency which depended on the salt concentration employed during haemolysis and on the concentration of the DNA itself. The entrapped DNA was largely protected from digestion by deoxyribonuclease I and could be recovered intact by phenol extraction. Erythrocyte ghosts containing 32P-adenovirus type 2 DNA were fused with KB cells during brief treatment with polyethylene glycol (PEG). Following fusion, counts equivalent to an average of 25 molecules of labelled DNA were micro-injected and transported to each K B cell nucleus. Less than ¼ as many DNA counts were recovered from nuclei when PEG treatment was omitted. A direct immunofluorescent assay demonstrated virus replication in some cells following their fusion with DNA-containing ghosts. The efficiency of transfection was considerably lower than that expected from the large number of successfully micro-injected DNA molecules. This suggests that most of the micro-injected DNA molecules were degraded before a successful infection could be completed.

This publication has 0 references indexed in Scilit: