Knockout lines for ProDH1 and ProDH2 and T-DNA excision during splicing. A: Graphic representation of the exon-intron structure of the ProDH1 and ProDH2 genes with the position of T-DNA insertions in the analysed mutant lines (see material and methods section). Arrows with letters indicate the binding sites of primers used for PCR analyses in B and C. B: PCR on genomic DNA with two primer pairs, one for RbohD (At5g47910 as internal control, fragment size 1272 bp) and one ProDH1 or ProDH2 specific primer pair. Absence of ProDH specific PCR products demonstrates that all mutant plants were homozygous for the respective T-DNA insertion. C: RT-PCR analysis of ProDH1 and ProDH2 expression in the mutant lines demonstrates the presence of native transcripts in pdh1-4 and pdh2-2. PCR reactions with cloned cDNAs and genomic DNA (leftmost three lanes) demonstrate specificity of the PCR products. cDNAs for C and genomic DNA for B were obtained from the same samples, a slight contamination of the RNA samples with genomic DNA caused the additional amplification of intron-containing PCR products in the RT-PCRs.