I wonder if I infected two cells with IAV using two different MOI, one of them 1 and the other is 10. Now I wanted to compare the expression level of one gene in both cells. I think this is not right to compare. has anyone any suggestion to surmount or overcome this mistake without repeating the experiment. Any suggestions?
I think that you should compare the gene expresion of each infected sample with a mock infected cels (no virus) and I think that is correct.
Might be, what I did that I compare the gene level in each cell with B actin as a control and this will done separately in each cell then I can have the reference gene and finally I can say that this gene is higher in this cell than the other upon infection , is this is right
You should keep in mind that an MOI of 1 means that many cells will be uninfected (if you use a poison distribution, you get 1/3 of cells uninfected at this MOI), and hence will not have any change in expression. MOI of 3 should give you almost all cells infected.
Hence, you should plan your experiment according to what you want to see. If you are just looking if viral infection changes a gene, use an MOI of at least 3 and you should see an effect. To push the system further, you can use a much higher MOI to see if higher number of viruses infecting a cells makes a difference, and in both cases, compare the results to an uninfected control to know that your change comes from the infection.
Good but my question is i want to say that this gene got upregulated or down in one cell but not the other. here I have to fix the MOI since different MOI produce different results
My question is how to overcome my problem in which I infected two cell line with different MOI of the same virus and I want to say that this gene is expressed differantially in the two cell system. Is my different MOI affect the results. My sensation is yes but how can I overcome without repeating
I think you probably want to repeat it to be sure, and compare it to uninfected. Different MOI can influence things like interferon activation/blocking, and kinetics of cell expression changes, especially if in one condition you have many cells that are uninfected.
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