| Property | Value |
|---|---|
| BioDare ID | 3488 |
| Author | Harriet G. McWatters |
| Institution | University of Oxford |
| License | CC_BY |
Literature data from: 'Defining the robust behaviour of the plant clock gene circuit with absolute RNA timeseries and open infrastructure.' by: Harriet G. McWatters.
data are col0 8 d seedlings grown on sucrose in either LD 12:12 at 20 oC or WC 27:12 in LL.
Gene expression normalised on btub4 control RNA in same sample at same time. Each datapoint is mean of two biological replicates (each with three technical replicates).
Harriet McWatters study to compare clock gene expression with light and temperature entrainment, published in Flis et al. 2015
Experiment by Harriet McWatters lab in Oxford, perhaps ca. 2007. Published in Flis et al. RS Open Biology 2015. Prep details From Knight, Thomson and McWatters PP 2008: "seedlings were grown on plates containing 1× MS salts with 0.8% to 1% agar and, where indicated, supplemented with 3% (w/v) Suc. In all cases, media pH was corrected to pH 5.8. Unless otherwise stated, seeds were surface-sterilized before being stratified in the dark at 4°C for 48 h prior to transfer to the growth chamber. Plants were grown in Sanyo MLR-350 growth chambers at a constant temperature (20°C). The light level during photoperiods or constant light was 50 μmol m−2 s−1."
From Knight et al. 2008: Samples of 20 to 30 seedlings were collected every 3 h and snap-frozen in liquid nitrogen; the first sample was collected at subjective dawn on day 14, after 24 h in LL. Similarly, for analysis of flowering time gene expression, seedlings were grown on plates as described above in LD 16:8 photoperiods for 13 d and sampled every 2 h on day 14 in a free-running cycle. RNA was extracted (RNeasy kit, Qiagen [74904] with additional DNAse digestion) from each sample and cDNA synthesized (Taq-Man, Applied Biosystems [N808-0234] reverse transcriptase kit) for each time point.
RT-PCR (SYBR Green PCR Master mix (Applied Biosystems [4309155]) using an Applied Biosystems Prism-7300.)
Literature data from: '' by: .
From Knight et al. 2008: Samples of 20 to 30 seedlings were collected every 3 h and snap-frozen in liquid nitrogen; the first sample was collected at subjective dawn on day 14, after 24 h in LL. Similarly, for analysis of flowering time gene expression, seedlings were grown on plates as described above in LD 16:8 photoperiods for 13 d and sampled every 2 h on day 14 in a free-running cycle. RNA was extracted (RNeasy kit, Qiagen [74904] with additional DNAse digestion) from each sample and cDNA synthesized (Taq-Man, Applied Biosystems [N808-0234] reverse transcriptase kit) for each time point. Real-time reverse transcription (RT)-PCR was carried out in triplicate with SYBR Green PCR Master mix (Applied Biosystems [4309155]) using an Applied Biosystems Prism-7300. Levels of specific mRNA and βTUBULIN4 (βTUB4) controls were calculated by the standard curve method (Applied Biosystems user bulletin 2); the relative expression (arbitrary units) of each gene of interest was obtained by dividing by contemporaneous βTUB4 expression. No-RT and no-template controls were included as negative controls for each set of reactions. Two or three independent biological repeats gave similar results.
Growth on Agar 1.5% MS 1 3% Sucrose for 7.0 days (LD12:12). Actually less agar (0.8-1%) according to the paper
| Type | Duration (days) | Cycle (h) | Start | Duration | Spectrum | Source | Intensity |
|---|---|---|---|---|---|---|---|
| diurnal light | 7 | 24 | 0:00 | 12:00 | white | tube | 50 |
| Type | Duration (days) | Cycle (h) | Base (°C) | Warm (°C) | Warm start | Warm duration |
|---|---|---|---|---|---|---|
| constant temperature | 7 | 24 | 20 | -- | -- | -- |
Growth on Agar 1.5% MS 1 3% Sucrose for 7.0 days (WC 27oC:12oC in LL). Actually less agar (0.8-1%) according to the paper
| Type | Duration (days) | Cycle (h) | Start | Duration | Spectrum | Source | Intensity |
|---|---|---|---|---|---|---|---|
| constant light | 7 | 24 | 0:00 | 24:00 | white | tube | 50 |
| Type | Duration (days) | Cycle (h) | Base (°C) | Warm (°C) | Warm start | Warm duration |
|---|---|---|---|---|---|---|
| diurnal temp | 7 | 24 | 12 | 27 | 0:00 | 12:00 |
Growth on Agar 1.5% MS 1 3% Sucrose for 1.0 days (LD 12:12). same as entraining condition
| Type | Duration (days) | Cycle (h) | Start | Duration | Spectrum | Source | Intensity |
|---|---|---|---|---|---|---|---|
| diurnal light | 1 | 24 | 0:00 | 12:00 | white | tube | 50 |
| Type | Duration (days) | Cycle (h) | Base (°C) | Warm (°C) | Warm start | Warm duration |
|---|---|---|---|---|---|---|
| constant temperature | 1 | 24 | 20 | -- | -- | -- |
Growth on Agar 1.5% MS 1 3% Sucrose for 1.0 days (WC 27oC:12oC in LL). same as entraining condition
| Type | Duration (days) | Cycle (h) | Start | Duration | Spectrum | Source | Intensity |
|---|---|---|---|---|---|---|---|
| constant light | 1 | 24 | 0:00 | 24:00 | white | tube | 50 |
| Type | Duration (days) | Cycle (h) | Base (°C) | Warm (°C) | Warm start | Warm duration |
|---|---|---|---|---|---|---|
| diurnal temp | 1 | 24 | 12 | 27 | 0:00 | 12:00 |