A series of C4N hairpin RNAs bearing anticodon nucleotides at the 5' ends and a discriminator base and the sequence CCA at the 3' ends was constructed by an in vitro transcription system using T7 RNA polymerase. These RNAs were aminoacylated specifically with their cognate amino acids by reaction with aminoacyl-adenylates in the presence of a dipeptide, valyl-aspartic acid, suggesting that such hairpin RNAs are able to play the role of the present-day tRNA and that valyl-aspartic acid can perform the function of the present-day aminoacyl-tRNA synthetase as a catalyst in the aminoacylation reaction. These results should provide a useful clue to elucidating the origin of the genetic code.