Figure 4.

Reduced levels of γ1 circle transcripts (CT) but unaltered levels of γ1 germline transcripts (GLT) in activation-induced deaminase (AID) heterozygotes indicate that the defect in heterozygotes is downstream of transcription of switch regions. Resting/naïve B cells were prepared from AID wild-type mice (n= 3; WT) and heterozygotes (n= 5; Het) on the MRL/lpr background. B cells with the same genotypes were pooled and treated with lipopolysaccharide plus interleukin-4 (LPS + IL-4) for 24 hr and 48 hr. Cells were lysed in TRIzol for RNA preparation. Complementary DNA was synthesized and used at different dilutions to determine γ1 circle transcripts and germline transcripts by reverse transcription–polymerase chain reaction (PCR). The PCR products were visualized in agarose gel and quantified by imagequant software. The reduction in circle transcripts was observed in at least two separate experiments. WT, wild-type; H, heterozygous.