Differential mRNA display of T-cell subsets derived from the anti-CD3–inducible T-cell differentiation system. (A) Arrested double-negative DNIII thymocytes from untreated as well as DNIV immature single-positive (ISP) and double-positive (DP) thymocytes from anti-CD3ϵ–treated Rag ko mice were sorted in doublet, and total RNA was used for differential mRNA display. Zap-70 mRNA was identified twice with the 2 arbitrary 5′ primers AP02 (200 bp, left) and AP11 (449 bp, right) in combination with T11G primers. (B) Germ-line TCRα transcripts were found with the primer set AP16 and T11G (264 bp). Northern blot of total RNA from thymi of the indicated mice (right panels) confirmed the onset of germ-line TCRα transcription in anti-CD3ϵ–induced Rag ko thymocytes. (C) With 2 different primer sets, AP03/T11G (left) and AP04/T11G (right), a single differentially expressed cDNA species was identified independently. (D) Semiquantitative PCR with gene-specific primers verified the differential expression of Gimap4; depicted are 40, 30, and 20 cycles from left to right. (E) Northern blot of RNA from thymi of anti-CD3ϵ–treated or nontreated Rag ko mice confirmed quantitatively the expression and the CD3-dependent induction of Gimap4. A labeled cDNA probe revealed a single message of 1.8 kb.