the vector is pBV-luc with myc gene insert, what would be an easy way to check if the vector works?
Hi I am actually looking for the myc-reporter and noticed your question.
What have you tried so far?
Transfect U2OS cells that express low levels of endogenous MYC and stimulate with doxycycline.
Transfect in a colon cancer cell line and it should be expressed relatively highly.
If any reply please include me: [email protected]
I just was thinking in sequencing the vector or in the digestion as well. But I was wordering if there was an experimental aproach firts. Thank you very much!
MYC stimulates the expression of many genes, but typically with low fold induction, ca. 1.5x-3x in reporter assays. The ODC1 promoter is a reliable positive control to test whether your assay actually works.
As the levels of activation by MYC are typically low, you have to very carefully control the conditions of the experiment to get consistent results, like keeping the cell numbers constant, same medium and serum, cell line with low level of endogenous MYC activity... I also like U2OS for reporter assays, they are quite easy to transfect and in assays using miRNA, they tend to give consistent results between experiments.
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