Figure 1.

Selective silencing of A. thaliana 45S rRNA gene subtypes. (A) 45S rRNA genes are repeated in long tandem arrays at NORs located on chromosomes 2 and 4. Each rRNA gene repeat includes a 45S pre-rRNA transcription unit and an intergenic spacer (IGS) that includes the gene promoter. The 5′ and 3′ external transcribed spacer sequences (5′ ETS and 3′ ETS) and internal transcribed spacers (ITS1 and ITS2) are removed during pre-RNA processing. Within the 3′ ETS, a variable region defines rRNA gene types Variant 1 (VAR1) through VAR4. (B) Primers flanking the 3′ ETS variable region yield PCR products that define rRNA gene types VAR1, VAR2, VAR3, and VAR4. The gel image at the right shows PCR products upon amplification of genomic DNA or cDNA of leaf tissue RNA, comparing wild-type and an hda6-null mutant. PCR primer sequences are provided in Supplemental Figure S1. (C) Locations of nucleotide polymorphisms that define rRNA gene subtypes. Positions of polymorphic nucleotides are numbered relative to the 45S rRNA gene reference sequence provided in Supplemental Figure S2. Single-nucleotide polymorphism names begin with the nucleotide of the reference sequence, followed by the position in the reference sequence, and ending with the alternative nucleotide at that position. 6978+CAT is a trinucleotide insertion (CAT) relative to the reference sequence. The relative frequencies of the alternative polymorphic nucleotides is provided in Supplemental Figure S3. (D) Differential expression of rRNA gene subtypes defined by sequence polymorphisms. The diagrams at the left of the gel images show the positions of PCR primers flanking the polymorphic site and the natural or derived restriction endonuclease recognition site that results from the polymorphism. Gel images show PCR products obtained using genomic DNA or RT–PCR in the wild type (Col-0) or hda6 mutant (in the Col-0 genetic background) following digestion with the relevant restriction endonuclease. Uncut controls are also shown in each case.