Fig. 2: Transcriptional and epigenetic status of Igh alleles with Eµ substituted by EF1α promoter.

a Transcriptional analysis of WT, Eµ-deficient, and EF1α promoter replacement Igh alleles. Eµ generates bi-directional (sense and antisense) RNA. Eµ was replaced by the EF1α promoter in two different orientations. EF1α^For directs transcription toward the Cµ (sense orientation), whereas EF1α^Rev directs transcription toward D_H-J_H region (antisense orientation). RT-qPCR was performed using primer pairs S1–S5 to assess the transcription profiles of various Igh alleles as indicated. Transcription levels relative to WT are shown as the bar graphs. The data are presented as mean ± SEM from three independent experiments. Results from two different EF1α^For and EF1α^Rev clones (#1 and #2) are shown. Pair primers P1–P4, P2–P4, and P3–P4 were used for DJ_H rearrangement analysis of WT, Eµ-deficient, and EF1α promoter replacement alleles (see Fig. 3c). The Experiment was independently repeated three times with similar results. Source data are provided as a Source Data file. b Transcriptional and epigenetic features of Eµ^−/− and EF1α promoter replacement Igh alleles. Directional RNA-seq and ChIP-seq analysis of Eµ manipulated cell lines are indicated. The 3′ Igh domain extending from IGCR1 to the 3′CBE is displayed. IGCR1, DQ52-Cµ and Eμ loci are highlighted by black, red and green rectangles, respectively. β-actin locus (chr5:142,890,001-142,940,000) was used as a control (right). Experiment was independently repeated twice with similar results. See also Supplementary Fig. 2, Data 1, and 6.