A method and apparatus for transmitting a Reference Signal (RS) during retransmission at a User Equipment (UE) in a multi-antenna wireless communication system are disclosed. The RS transmission method includes receiving an uplink grant from a Base Station (BS), initially transmitting data and an RS corresponding to each layer based on the uplink grant, receiving ACKnowledgment/Negative ACKnowledgment (ACK/NACK) information for the initially transmitted data, and retransmitting the RS corresponding to each layer and the data according to the ACK/NACK information. The retransmission includes initializing a cyclic shift value for the RS corresponding to each layer according to a cyclic shift field for RSs included in the uplink grant.