Why do genes, encoding the same proteins and in the same conditions, have different expression?
If I understand your question and graph correctly, your Y-axis is log(x/REF), where REF is some external standard. Your &quot;Ref&quot; on the x-axis you expect to be the same as REF, so that log(Ref/REF) &quot;should be&quot; zero, but you find it is not.
However, it looks like the mean(log(Ref/REF)) is still approximately zero. This is what I would expect: values that deviate a bit from perfectly 0, but having an average around 0 (and this average should approach 0 the more samples you have per the law of large numbers). You don't expect to have zero variance in data like this, which is why you repeat the experiment a number of times. You'll notice you have variance in all of your categories.