Transfection of hepatic genes into adult rat hepoatocytes in primary culture and their tissue‐specific expression
- 3 March 1989
- journal article
- research article
- Published by Wiley in European Journal of Biochemistry
- Vol. 180 (2), 289-294
- https://doi.org/10.1111/j.1432-1033.1989.tb14646.x
Abstract
We describe in this paper a method for studying transient gene expression in a primary culture of adult rat hepatocytes. After isolation by collagenase perfusion, hepatocytes in a monolayer were transfected with foreign DNA by the calcium phosphate precipitation technique during the first 24 hours after plating. When they were transfected with a plasmid containing the gene for chloramphenicol acetyltransferase driven by the early promoter of simian virus 40, hepatocytes reproducibly expressed high levels of chloramphenical acetyltransferase (CAT); this transient expression was much higher than that obtained with the rat hepatoma cell line H411. Different medium conditions have been tested; an optimal level of CAT activity can be obtained using a serum-free, hormonally defined medium. Using these techniques, we have investigated the expression of liver-specific genes transferred into hepatocytes. We show that the L-pyruvate kinase promoter is active in these hepatocytes while it is silent in fibroblasts. Moreover, the use of serum-free medium may allow investigation of the role of hormones and nutrients in cells which respond normally to these effectors.This publication has 28 references indexed in Scilit:
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