title: Large macromolecules can be introduced into cultured mammalian cells using erythrocyte membrane vesicles.
abstract: Plasmid 6.4 kbp DNA, 14 kbp DNA, lambda phage particles, all of which contained herpes simplex virus type 1 (HSV-1) thymidine kinase (TK) gene, or IgM molecules, were mixed with erythrocyte membranes and treated with neutral detergent. The transparent mixture was diluted with phosphate-buffered saline (PBS), followed by centrifugation to collect membrane vesicles containing the large macromolecules. 10-15% of 6.4 kbp, 3% of 14 kbp, 4-7% of the lambda phage particles and 14.5% of IgM were trapped within erythrocyte membrane vesicles. The membrane vesicles containing these molecules were fused with L cells, or rat F2408#20 cells, both of which are deficient in thymidine kinase activity. In each case, transformants were obtained. 2 X 10(5) - 7 X 10(5) phage PFU or 1.5 X 10(6) - 8 X 10(7) DNA molecules were required to obtain one transformant from L cells, but 2-3 X 10(7) phage PFU or 2 X 10(9) - 1 X 10(10) DNA molecules were required for one transformant from rat cells. Number of colonies which transiently expressed TK genes in L cells was also determined by autoradiography. The ratio of stable transformants to colonies positive for transient expression in cells treated with low doses of DNA or lambda phage was 46-68%. The transformation efficiency of human fibroblast cells by pSV2-gpt DNA trapped in erythrocyte membrane vesicles was less than that of L cells by HSV-TK DNA, but almost the same as that of rat cells by HSV-TK DNA.
authors.forename: H
authors.initials: H
authors.lastname: Sugawa
authors.forename: T
authors.initials: T
authors.lastname: Uchida
authors.forename: Y
authors.initials: Y
authors.lastname: Yoneda
authors.forename: M
authors.initials: M
authors.lastname: Ishiura
authors.forename: Y
authors.initials: Y
authors.lastname: Okada
mesh_terms: D000818:Animals; D010582:Bacteriophage lambda; D002478:Cells, Cultured; D003584:Cytological Techniques; D004910:Erythrocyte Membrane; D006801:Humans; D007075:Immunoglobulin M; D007739:L Cells; D008561:Membrane Fusion; D051379:Mice; D010957:Plasmids; D051381:Rats; D013937:Thymidine Kinase / Q000235:genetics
chemical_list: D007075:Immunoglobulin M; D013937:Thymidine Kinase