Supplemental Data

Files in this Data Supplement:

• Supplemental Figure 1 - Supplemental Figure 1. Identification and Molecular Characterization of ahk2 and ahk3 Mutant Alleles. (A) Structures of the AHK2 and AHK3 genes and insertion sites of the T-DNAs in the ahk2 and ahk3 mutant alleles used in this study. Thick lines represent exons. T-DNA insertions are indicated as triangles and are not drawn to scale. (B) Sequence at the genome-T-DNA junction sites in the ahk2 and ahk3 mutant alleles. The sequences of the genes are shown on a gray background, the sequence of the T-DNA is printed in bold, and sequences of unknown origin found in two of the insertion lines are underlined. Numbers above the sequence indicate the base pair position relative to the predicted translational start codon of the respective gene. (C) RT-PCR analysis of AHK2 and AHK3 transcripts. Primers specific for the AHK2 transcript and the AHK3 transcript were used to analyze the ahk2-5 and ahk3-7 mutants, respectively. The ahk2-5, ahk3-7 double mutant was analyzed with primers for both genes. The upper band corresponds in all cases to the actin2 transcript, which was used as a positive control. M, 1 kb ladder, the size in bp is indicated on the right; A, B, primer combinations to detect the AHK2 and AHK3 transcripts were used, respectively.
• Supplemental Figure 2 - Supplemental Figure 2. The Cytokinin Sensitivity of Cytokinin Receptor Mutants is Decreased in Shoots and Roots. (A) Reduced callus formation and regeneration capacity of ahk2 and ahk3 mutant hypocotyls on media containing different auxin (NAA) and cytokinin (2iP; isopentenyl adenine) concentrations. The hypocotyl segments of seven day old plants were cut off and cultured for 28 days on different hormone media as shown. Representative samples for each tested concentration were placed together prior to photography. (B) Enhanced growth, leaf greening and flowering capacity of ahk2 ahk3 double mutants compared to wild type (WT) and single mutants on medium supplemented with cytokinin (0.1 mg/l BA). Photo was taken 28 DAG (days after germination). (C) Elongation of the primary root of wild type (WT) and mutant plants between 4 DAG and 7 DAG on media containing different concentrations of BA. Root elongation on cytokinin-free medium was set at 100% for each genotype. Root elongation on this medium was for WT, 1.06 ±0.17 cm; ahk2, 1.05 ±0.18 cm; ahk3, 1.06 ±0.16 cm; ahk2 ahk3, 1.42 &plusmn0.38 cm; ahk2 cre1, 0.86 ±0.27 cm; ahk3 cre1, 1.21 ±0.25 cm) and ahk2 ahk3 cre1, 1.24 &plusmn0.32 cm. Error bars represent SE (n=15). (D) Elongation of the primary roots of ahk2 ahk3 mutants (the two seedlings on the right) compared to WT on MS medium containing 0.1 mg/l BA 21 DAG.