| Property | Value |
|---|---|
| BioDare ID | 13790739219191 |
| Author | Niwa Y. |
| Institution | Nagoya University |
| License | CC_BY |
In Arabidopsis thaliana, the flowering time is regulated through the circadian clock that measures day-length and modulates the photoperiodic CO–FT output pathway in accordance with the external coincidence model. Nevertheless, the genetic linkages between the major clock associated TOC1, CCA1 and LHY genes and the canonical CO–FT flowering pathway are less clear. By employing a set of mutants including an extremely early flowering toc1 cca1 lhy triple mutant, here we showed that CCA1 and LHY act redundantly as negative regulators of the photoperiodic flowering pathway. The partly redundant CCA1/LHY functions are largely, but not absolutely, dependent on the upstream TOC1 gene that serves as an activator. The results of examination with reference to the expression profiles of CO and FT in the mutants indicated that this clock circuitry is indeed linked to the CO–FT output pathway, if not exclusively. For this linkage, the phase control of certain flowering-associated genes, GI, CDF1 and FKF1, appears to be crucial. Furthermore, the genetic linkage between TOC1 and CCA1/LHY is compatible with the negative and positive feedback loop, which is currently believed to be a core of the circadian clock. The results of this study suggested that the circadian clock might open an exit for a photoperiodic output pathway during the daytime. In the context of the current clock model, these results will be discussed in connection with the previous finding that the same clock might open an exit for the early photomorphogenic output pathway during the night-time.
They were harvested (3 or 3.5 h intervals) to quantify the mRNA of genes involved in flowering regulation. Total RNA was purified by an RNeasy kit (Qiagen, Valencia, CA, USA). To synthesize cDNA, 1 mg of each RNA was reverse transcribed with ReverTea Ace (TOYOBO, Osaka, Japan) and oligo(dT) primer. The synthesized cDNAs were amplified with iQ SYBR Green supermix (Bio-Rad Laboratories, Hercules, CA, USA) and a primer set using a MiniOpticon real-time PCR system (Bio-Rad Laboratories). The primer sets used in this study were, for CO, 5'-CTACAACGACAATGGTTCCATTAAC-3' and 5'-CAGGGTCAGGTTGTTGC-3'; for GI, 5'-GGGTAAATATGCTGCTGGAGA-3' and 5'-CAGTATGACACCAGCTCCATT-3'; for CDF1, 5'-TTTCCCGACGGTTTTAGAGG-3' and 5'-CCATGCTGTTGCATCTTGGA-3'; and for FKF1, 5'-GAAGTCTTCACTGGCTATCG-3' and 5'-GATCAACCAATGGGTGACG-3'. Primer sets for APX3 and FT were described by Hazen et al. (2005) and Endo et al. (2005), respectively. The following standard thermal cycling program was used for all PCRs: 958C for 120 s, 40 cycles of 958C for 10 s, and 608C for 60 s. Data were analyzed using Opticon Monitor version 3.1 software (Bio-Rad Laboratories).
RT-PCR ()
Growth on MS plates 1% sucrose for 10.0 days (Niwa2007).
| Type | Duration (days) | Cycle (h) | Start | Duration | Spectrum | Source | Intensity |
|---|---|---|---|---|---|---|---|
| diurnal light | 10 | 24 | 0:00 | 10:00 | white | LED | 120-150 |
| Type | Duration (days) | Cycle (h) | Base (°C) | Warm (°C) | Warm start | Warm duration |
|---|---|---|---|---|---|---|
| constant temperature | 10 | 24 | 22 | -- | -- | -- |
Growth on MS plates 1% sucrose for 1.0 days (Niwa2007).
| Type | Duration (days) | Cycle (h) | Start | Duration | Spectrum | Source | Intensity |
|---|---|---|---|---|---|---|---|
| diurnal light | 1 | 24 | 0:00 | 10:00 | white | LED | 120-150 |
| Type | Duration (days) | Cycle (h) | Base (°C) | Warm (°C) | Warm start | Warm duration |
|---|---|---|---|---|---|---|
| constant temperature | 1 | 24 | 22 | -- | -- | -- |