Transfection of liver in vivo by biolistic particle delivery |
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Authors: | Roongsiri Muangmoonchai Siew Cheng Wong Despina Smirlis Ian R. Phillips Elizabeth A. Shephard |
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Affiliation: | (1) Department of Biochemistry and Molecular Biology, University College London, Gower Street, WC1E 6BT London, UK;(2) School of Biological Sciences, Queen Mary, University of London, Mile End Road, E1 4NS London, UK;(3) Present address: Department of Clinical Chemistry, Faculty of Associated Medical Sciences, Chiang Mai University, 50200 Chiang Mai, Thailand;(4) Present address: Institute of Molecular and Cell Biology, National University of Singapore, 30 Medical Drive, 117609 Singapore, Republic of Singapore;(5) Present address: Department of Biochemistry, Molecular Parasitology Laboratory, Hellenic Pasteur Institute, Athens, Greece |
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Abstract: | We describe the delivery of reporter gene constructs to rat liver through the use of the Helios Gene Gun system. The effectiveness of this transfection method is illustrated by describing its use for determining in vivo the role of a DNA element that regulates cytochrome P450 2B1 (CYP2B1) gene expression in response to xenobiotics. DNA was delivered to the liver of an anesthetized animal via DNA-coated gold microcarriers. The highest level of reporter gene expression was obtained about six hours posttransfection; however, at this time endogenous CYP2B1 mRNA is transiently induced by the anesthetic treatment. The optimal time for investigating expression of a reporter gene under the control of CYP2B1 regulatory elements was 24 h after transfection, by which time the inductive effect of the anesthetic had ceased. Reporter gene expression subsequently declined rapidly to a low level by 48 h. In the transfected liver the heterologous SV40 promoter was about eight-fold stronger than the minimal CYP2B1 promoter. However, when attached to the phenobarbital response element both promoters give the same fold-induction of reporter activity in response to phenobarbital. |
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Keywords: | Transfection liver gene regulation gene gun biolistic particle delivery cytochrome P450 |
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