Definitely Senju DNA was what Kabuto added to his body.
Based on his reaction, I'd say that this is the FIRST time he has ever had Senju DNA in his original body. However, most people on the base will prlly tell you that Kabuto added MORE senju DNA to his body. Basically most people on the base think that Rinnegan requires Senju DNA so he must have had it previously.
Personally, I think Madara never added senju DNA to himself, only Tobi, and that's why he was looking under his shirt; to see if Kabuto had added it. Therefore, I believe that Senju DNA is not required to unlock Rinnegan.