A clone has been selected from a cDNA library previously constructed from phenobarbital pre-treated rat liver polysomal poly(A)+ RNA, which was reverse transcribed. The double-stranded cDNA was inserted by GC homopolymeric tailing in the Pst I site of pAT 153, and further cloned in E. coli HB101. This clone, called 2A9, corresponds to a mRNA whose concentration is increased five fold 16 h after phenobarbital treatment. Its length is 1200 nucleotides as revealed by RNA dot and Northern blot analysis respectively. The two strands of a 450 bp fragment from the 2A9 580 bp double-stranded cDNA insert have been sequenced and proven to correspond to alpha 2u globulin mRNA. It shows one single bp difference from the sequence previously published by Unterman et al. (1981, PNAS, 78, 3478). Thus, alpha 2u globulin, a hormone regulated gene product, is inducible by phenobarbital.