Assays

The list of python packages used was obtained by typing inside the Docker image

pip list -- format==columns > python_packages_pip_installed.txt

This list the version of packages installed as we have observed issues related to the use of the most current version of some python packaged for example scipy

Maps of cloned genes for rescuing selected clock mutants

The promoter regions for clock genes that present a ChIP-seq signal were extracted from TAIR10 using costume python scripts using the gene list for Kamioka et al CCA1 or Daphne Ezer et al for LUX. The promoter was considered from the TSS of the gene until the annotated end of the upstream gene. Then, this region was scanned using the Energy Matrix derived using EMA working as a classifier for bound or unbound. After classification the calibrated PBM data calibrated using in vitro data was used ...

The reporter fusion constructs expressing clock proteins fused to NanoLUC or firefly FLUC were transformed into the cognate, clock-mutant host plants. Each host also contained a transcriptional FLUC fusion that was used to score the circadian period of each transgenic line in constant light. Transformants that expressed a functionally normal level of clock protein were selected by choosing lines that complemented the mutant's period defect back close to the wild type period. Note that the reporters ...

This section contains the links to the tools used for reproducing the computational results presented in Urquiza-Garcia et al. 2022. This is required in particular because the SloppyCell model optimisation software is at some risk. Using Docker we can assure persistence for the computational environment that allows you to run SloppyCell.

The associated git repository can be found in https://hub.docker.com/r/uurquiza/urquiza2019a_tellurium_sloppycell/tags which can be cloned.

The docker image can ...

Analysis for inferring the number of molecules of clock proteins using recombinant NanoLUC

This script take the scaling paramters using synthetic protein data updates model paramaters

Seedlings of the transgenic lines complemented with CCA1-NL and TOC1-NL were tested under 12L:12D cycles followed by constant light, to test how well the reporter signal in living plants reflected the expected patterns of protein expression. One example is linked below, from the BioDare2 repository record, because FAIRDOMHub's Data File is not accepting these URLs.

BioDare2 ID 11391; Plate reader experiment CCA1 TOC1 NanoLUC; permalink: https://biodare2.ed.ac.uk/experiment/11391

Jupyter lab notebook that contains the models and data that for predicting protein levels based on mRNA data from TiMet projecto

RNA timeseries data for Arabidopsis Col wild-type plants and clock mutants, as separate mean and SD files. The raw data is available on BioDare.ed.ac.uk, and is linked as 'Attribution' from elsewhere on FAIRDOMHub.

Protein time series for clock proteins colected from the literature. Protein expression profiles were derived from images of western blot or or from plots that the orignal authors derived from quantitative western blots

Insertion of events that rescued mutant phenotypes were selected for performing absolute quantification using calibration curves of recombinant purified MBP-NanoLUC-3Flag-10his. Seeds were sterilised with 5% houshold bleach for 10 min and washed three time with deionised water. The seeds were then put for stratifyication at 4ºC in darkenss for 48 hours in 1.5 ml polyproplyen tubes in dionised water. After 48 hours seeds wered plated on ROBUST agar (1/2 MS salts, 1.2% Agar pH 5.8 ajudsted with ...

Leaf number at flowering data from literature for prr7 prr9 and Col wild-type plants under long photoperiods and short photoperiods

Seedling hypocotyl data from literature for prr7 prr9 and Col wild-type plants under various photoperiods

Biomass and metabolites in Col0 (WT) and prr7prr9, with and without exogenous gibberellins (GA)

Luciferase reporter gene assay for circadian period of seedlings in constant light, for Col0 (WT) and prr7prr9, with and without exogenous gibberellins (GA). Supplementary Figure 11f in Chew et al., _in Silico _Plants.

Raw and processed data, together with circadian period analysis and summary statistics, are available from BioDare.ed.ac.uk: choose https://biodare.ed.ac.uk/experiment ("Browse Public Resources" on the Login screen), then you can link to https://biodare.ed.ac.uk/robust/ShowExperiment.action?experimentId=3838, ...

Biomass, leaf number and gas exchange data for Col0 (WT), prr7prr9, and lsf1, compiled from four studies: L&H1-3 and the 'no GA' controls of Gibberellins 1.

Follow-up to the validation experiments on FMv2, testing candidate mechanisms for high malate and fumarate accumulation in the Arabidopsis double mutant prr7prr9 and its parent accession Col.

In this study, 14CO2 labelling was used to test the rate of carbon assimilation in the dark at the end of the subjective night (starting about ZT21), which is indicative of PEPC activity in forming malate, and the subsequent partitioning of this labelled C into various cellular fractions. The short-period ...

Follow-up to the validation experiments on FMv2, testing candidate mechanisms for high malate and fumarate accumulation in the Arabidopsis double mutant prr7prr9 and its parent accession Col.

In this study, thiamine vitamers were quantified to test whether the essential cofactor TDP had altered enzyme activities to affect the malate and fumarate levels, using existing plant samples harvested from am earlier L&H study.

Biomass (fresh mass, dry mass), leaf numbers, leaf area, gas exchange and 12 metabolites in Col0 (WT), prr7prr9, and pgm at days 29 and 35, presented in the preprint/publication, with most data also for Col and lhycca1 at days 21/22/23, not analysed further.

We suggest that the lower carbon assimilation rate measured in lhycca1 (see gas exchange data) might allow a calibirated simulation in the FMv2 model in future to incorporate the indirect effects of nightly carbon starvation in this genotype ...

effects of 1% increase in each parameter, more detailed analysis of water content

correlations of starch mobilisation and fresh weight under single parameter changes

Comparison of simulated wild-type and prr7prr9 double mutant under 12L:12D cycles. Simulation with CVODE simulator via SBSI v1.5 framework.

Biomass (fresh mass, dry mass), leaf numbers, leaf area, gas exchange and 12 metabolites in Col0 (WT), prr7prr9, and lsf1 (presented in the preprint/paper) and pgm (not analysed further).

Simulation data from FMv2 calibrated for experiment L&H2, an experiment run at 18.5C instead of the 20.5C of the replicate and related studies. The Excel file includes the mean and SD of the relevant experimental data, and the figure panels.

Metabolite analysis in clock mutants: Col-0 parent and mutants gi-201, toc1-101 and prr7prr9; WS parent and lhy/cca1 double mutant. Plants grown in Golm and harvested at End of Day and End of Night, , assays 22 major metabolites. More detail on TiMet wiki if required. Heteroscedastic t-tests to highlight most significant changes, without multiple-testing correction.

RNA timeseries data from TiMet for clock genes in prr7 prr9 and Col wild-type plants under 12L:12D cycle and LL

Combination of multiple sub-models to form Framework Model version 2

Biomass, leaf number and metabolites in Col0 (WT), prr7, prr7prr9, and lsf1. Metabolite data from plants after 28 days of growth were analysed most (27 days 'end of night', 28 days 'end of day' and 'end of night'). The data file also includes data from 21 days of growth ('end of day' and 'end of night'), which is useful for comparison to early-flowering plants not tested here, such as the lhycca1 double mutant, that flower before 28 days, altering their physiology.