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. 2009 Aug;16(4):237–247. doi: 10.1093/dnares/dsp012

Figure 4.

Figure 4

Quantitative confirmation of cytokinin-induced expression of type-A RR genes in L. japonicus seedlings. (A) Cytokinin-induced expression of type-A RR genes in L. japonicus seedlings. Seedlings were grown MS gellan gum-plates under constant light conditions for 17 days, and then these seedlings were sprayed with cytokinin (20 µM t-zeatin in 0.02% DMSO). The seedlings were harvested immediately before and 0.5, 1.0, 3.0 h after the treatment and subjected to RNA preparation. A set of indicated RR transcripts were analyzed by means of semi-quantitative RT–PCR to measure the amounts of each transcript. The UBC transcript encoding ubiquitin-conjugating enzyme (chr1.LjT04O06.40, a homologue of Arabidopsis At5g25760) was used as an internal control, the type-B RRb4 transcript was a negative control, and the CKX3 encoding cytokinin oxidase enzyme (LjT02N03.150, a homologue of Arabidopsis CKX4) was a positive control. The results of reference samples treated with 0.02% DMSO for 1.0 h were also presented at the most right-hand side. The primer set, used for these PCR analyses, was listed in Supplementary Table S2. (B) The essentially the same experiments were repeated in a more quantitative manner by means of real-time quantitative RT–PCR. The experiments were replicated three times to obtain mean values with SD.