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VSVG-Retroviral Production Problem


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#1 kriemo

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Posted 27 March 2009 - 02:13 PM

Howdy All,
I have a problem.
I am trying to generate retrovirus using a GP-293 cell line, with the VSVG envelope protein, however the VSVG plasmid seems to be lethal to the cells. I perform cotransfection with my transfer plasmid (pMIGR), and a VSVG expression construct (pMDG.2), with lipofectamine. 24 hours post transfection there are >70% GFP positive cells. However the cells with GFP have become ballooned in morphology, and after 48 hours have started to detach. The non-GFP cell morphology are not changed and continue to divide normally.
I see this "ballooning" with two different pMIGR transfer plasmids, one with a pri-microRNA sequence insert, and one without any insert. I also see that i can transfect these transfer constructs without VSVG into the GP-293 cell line, and it does not cause any ballooning and subsequent detachment.
Is there a significant amount of GP-293T lethality associated with VSVG psuedotyped viral production?

Thanks Everyone,
Kent
(bioforum newbie :rolleyes:

#2 little mouse

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Posted 31 March 2009 - 07:25 AM

I would say it's normal to see ballooning of your cells. They start to produce viruses. They don't like that so much. However you should still have some cells 72 hours after transfection, getting round and slightly detaching.




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