Transfer of mink genes into mouse cells by means of isolated lipid-encapsulated nuclei

• Published in: Molecular & general genetics Vol. 201; no. 3; p. 487
• Main Authors: Sukoyan, M A, Belyaev, N D, Budker, V G, Gradov, A A, Pack, S D, Serov, O L
• Format: Journal Article
• Language: English
• Published: Germany 01.01.1985
• Subjects: Animals; Cell Line; Genes; Liposomes; Mice; Mink - genetics; Mutation; Nuclear Transfer Techniques; Thymidine Kinase - genetics; Transformation, Genetic
• ISSN: 0026-8925; 1432-1874
• DOI: 10.1007/BF00331344

A method for gene transfer by means of interphase nuclei encapsulated within lipid membranes was developed. The method was based on passage of interphase nuclei through a layer of organic solvents containing phospholipids. Evidence was obtained indicating that the nuclei become surrounded by a protective phospholipid membrane: measurements of bound labelled or non-labelled phospholipids; decrease in the permeability of lipid-encapsulated nuclei for high molecular compounds; visualization by direct electron microscopy. Lipid-encapsulated nuclei of mink fibroblasts were used for transformation of mutant mouse LMTK- cells (deficient for thymidine kinase). The frequency of occurrence of HAT-resistant colonies/recipient cell was 1.9 X 10(-5). Biochemical analysis of 14 independent clones demonstrated that they all contained TK1 of mink origin. Analysis of 15 other biochemical markers located on 12 of the mink chromosomes revealed the activities of mink galactokinase (a syntenic marker) in 5 transformed clones, and that of mink aconitase-1 (the marker of mink chromosome 12) in 1 clone. No cytogenetically visible donor chromosomes were identified in the transformed clones. Nine transformed clones were tested for the stability of the TK+ phenotype; of these, the phenotype was expressed stably in 3 and unstably in 6. The method suggested is similar to the gene transfer procedure using total DNA. Its advantage is in ensuring efficient gene transfer and donor DNA integrity.