help with luciefarse reporter assay,:( - p53 reporter vector versus expression plasmid (Apr/26/2008 )
Im trying to see how my gene of inerest ( gene X) influences p53 activity under different apoptosis stimuli conditions. I have expression plasmid for gene X, which is psuper CMV 6 plasmid, and for p53 activity p53 reporter luciferase vector. I have also obtained control trans. luciferase vector. I'm trying to keep constant amount of Renilla vector as well, and p53 reporter luciferase vector, and changes amounts of plasmid for gene X to optimazed assay. My question is, what should be the ratio between pSport 6 plasmid coding for gene x and p53 reporter vector. I'm new in that field and I would like to ask for help. Thank you vecry much for your time and help.
I don`t know about your topic but I guess you would try different ratios. As you said, you could keep p53 lucif reporter plasmid amount const and use different amounts of effector gene (pSuper CMV6-geneX) and fill up the DNA amount with pUC18 or pUC19. Then you could put the lucif activity versus the amount of geneX in a graph. A positive and negative control must be used as well. But as I said I have no clue about your topic. Are there any papers published about that or any experiments done in your lab??