科研之友
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摘要
Reporter genes are important tools for assessing vector pharmacology in vivo. Although useful, current systems are limited by (1) the need to generate a new vector for each different reporter, (2) the inability to package reporter genes in small vectors, and (3) variations in reporter gene feedback due to variations in cell-to-cell vector copy number. To circumvent these problems, we have used Cre recombinase as a "cat's paw'' to activate reporter genes embedded in transgenic mice. The small Cre gene was introduced into self-complementary adeno-associated viral (scAAV) vectors with limited packaging capacity. Injection of scAAV-Cre vectors into mice with loxP-inactivated luciferase enabled in vivo imaging distributions comparable to the signal observed after AAV-luciferase injection. When injected into mT/mG mice, AAV-Cre converted ubiquitous expression of red fluorescent protein (RFP) to green fluorescent protein (GFP) expression only where the vectors transduced cells. Injection into F1 hybrid luciferase and mT/mG mice enabled simultaneous three-reporter tracking. This system was able to discriminate cell-specific transduction in all organs tested, with particular usefulness for detecting AAV serotype-specific transduction in the liver, kidney, and muscle. Given that F1 mice bear exactly one copy of luciferase and one copy of RFP-GFP, each reporter gene is either "on'' or "off'' in a cell. The Cre system therefore provides a unique quantum method to quantify vector delivery that can be applied when vector capacity is limited.
- 出版日期2012-10
