A method of transmitting a reference signal in a mobile communication system is provided. The method includes determining at least three scrambling variables comprising a first scrambling variable, a second scrambling variable, and a third scrambling variable, wherein the first scrambling variable, the second scrambling variable, and the third scrambling variable are different from each other, determining whether a demodulation reference signal is for more than or equal to 3-layer transmission, generating, if the demodulation reference signal is for more than or equal to 3-layer transmission, the demodulation reference signal sequence using the third scrambling variable and transmitting the demodulation reference signal sequence through a corresponding antenna port.