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Position: Discussion forums > Bioscience biotechniques discussion forums > Molecular biology troubleshooting forum
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Non functional adenovirus
Posted by: remadel (IP Hidden, New member, 1)
Date: February 15, 2005 01:30PM
Hello all! I wanted to see if someone can help me. I have two adenoviral constructs that I made using the Stratagene pShuttle-CMV-IRES-hrGFP vector. When I have tried to amplify these adenoviruses in HEK293 cells I lose the GFP fluorescence. Fluorescence was present upon transfection of the adenoviral construct with lipofectamine, but when I harvest the virus once, and then infect again to amplify it I lose the GFP expression. I see no fluorescence!! I tried a western to see if my protein is present in HEK extracts but it is not there. What is going on? What do I need to do? I have made many other adenoviruses before and this never happened. Any comments?
Re: Non functional adenovirus
Posted by: hz (IP Hidden, Unregistered user, )
Date: March 9, 2005 02:33PM
Two possibilities:
1. recombination and packaging didn’t occur. 2. MOI is too low and 293 harvested too early. other possibilities?
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