Asset Details
Do you wish to reserve the book?
In vivo electroporation and ubiquitin promoter - a protocol for sustained gene expression in the lung
Hey, we have placed the reservation for you!
By the way, why not check out events that you can attend while you pick your title.
Oops! Something went wrong.
Looks like we were not able to place the reservation. Kindly try again later.
Are you sure you want to remove the book from the shelf?
In vivo electroporation and ubiquitin promoter - a protocol for sustained gene expression in the lung
Do you wish to request the book?
In vivo electroporation and ubiquitin promoter - a protocol for sustained gene expression in the lung
Please be aware that the book you have requested cannot be checked out. If you would like to checkout this book, you can reserve another copy
We have requested the book for you!
Your request is successful and it will be processed during the Library working hours. Please check the status of your request in My Requests.
Oops! Something went wrong.
Looks like we were not able to place your request. Kindly try again later.
Journal Article
In vivo electroporation and ubiquitin promoter - a protocol for sustained gene expression in the lung
2006
Overview
Background Gene therapy applications require safe and efficient methods for gene transfer. Present methods are restricted by low efficiency and short duration of transgene expression. In vivo electroporation, a physical method of gene transfer, has evolved as an efficient method in recent years. We present a protocol involving electroporation combined with a long‐acting promoter system for gene transfer to the lung. Methods The study was designed to evaluate electroporation‐mediated gene transfer to the lung and to analyze a promoter system that allows prolonged transgene expression. A volume of 250 µl of purified plasmid DNA suspended in water was instilled into the left lung of anesthetized rats, followed by left thoracotomy and electroporation of the exposed left lung. Plasmids pCiKlux and pUblux expressing luciferase under the control of the cytomegalovirus immediate‐early promoter/enhancer (CMV‐IEPE) or human polyubiquitin c (Ubc) promoter were used. Electroporation conditions were optimized with four pulses (200 V/cm, 20 ms at 1 Hz) using flat plate electrodes. The animals were sacrificed at different time points up to day 40, after gene transfer. Gene expression was detected and quantified by bioluminescent reporter imaging (BLI) and relative light units per milligram of protein (RLU/mg) was measured by luminometer for p.Pyralis luciferase and immunohistochemistry, using an anti‐luciferase antibody. Results Gene expression with the CMV‐IEPE promoter was highest 24 h after gene transfer (2932 ± 249.4 relative light units (RLU)/mg of total lung protein) and returned to baseline by day 3 (382 ± 318 RLU/mg of total lung protein); at day 5 no expression was detected, whereas gene expression under the Ubc promoter was detected up to day 40 (1989 ± 710 RLU/mg of total lung protein) with a peak at day 20 (2821 ± 2092 RLU/mg of total lung protein). Arterial blood gas (PaO2), histological assessment and cytokine measurements showed no significant toxicity neither at day 1 nor at day 40. Conclusions These results provide evidence that in vivo electroporation is a safe and effective tool for non‐viral gene delivery to the lungs. If this method is used in combination with a long‐acting promoter system, sustained transgene expression can be achieved. Copyright © 2006 John Wiley & Sons, Ltd.
Publisher
John Wiley & Sons, Ltd,Wiley Periodicals Inc
