Fig. 2.
Expression of T11 gene. (A) PCR amplification of T11 and T23 genes from the wild type C57BL/6 (B6) and Qa-1b−/− (KO) mouse genomic DNA. The T11 amplicon size was 628bp and the T23 amplicon size was 677bp. (B) Reverse-transcriptional PCR amplification of T11 and T23 transcripts from B6 mouse spleen and thymus. The amplicon sizes are 415 bp and 438 bp for T11 and T23 respectively. RT: reverse transcriptase. (C) Quantitative-PCR to determine the T11 transcription level in different tissues. The total RNA was extracted from Qa-1b−/− mice. The reference gene was RNA polymerase 2A (POLR2A). The ratio of T11 to POLR2A was calculated according to the Pfaffl method. The qPCR was done in triplicates (n=3) and the experiment was repeated twice with similar results. One of them was shown. (D) Quantitative-PCR to determine the T11 transcription level in different cell subpopulations of thymus and spleen. The total RNA was extracted from thymus and spleen of Qa-1b−/− mice. The qPCR was done as above, at least in triplicates. (E) Qa-1/T23 and T11 expression after stimulation. Splenocytes of B6 or Qa-1b−/− mice were stimulated with plate-bound anti-CD3/28 Ab. Surface expression of Qa-1 on B6 splenocytes was determined by FACS. The total RNA was extracted from unstimulated or stimulated B6 or Qa-1b−/− splenocytes. The qPCR for T23 used RNA from B6 or for T11 used RNA from Qa-1b−/− splenocytes, respectively. The qPCR was done as same as above, in triplicates (n= 3) and the experiment was repeated twice with similar results. One of surface staining of Qa-1was shown.