Question (2331)
Lack of GFP expression can occur even when delivery appears successful. Expression levels may be influenced by cell type, promoter activity, transduction efficiency, or cellular stress. In some cells, CMV or other strong promoters may be silenced, and high MOI can negatively impact cell health.
Basic troubleshooting steps include:
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Confirming successful delivery using an alternative marker or control construct.
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Assessing cell viability and morphology after transduction.
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Allowing sufficient time post-delivery for expression.
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Evaluating whether the promoter is active in your specific cell line.
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Verifying construct integrity and GFP coding sequence.
Because expression outcomes are highly system-dependent, optimization may be required for each experimental setup.