In many species, temperature-sensitive phenotypic plasticity (i.e., an individual's phenotypic response to temperature) displays a positive correlation with latitude, a pattern presumed to reflect local adaptation. This geographic pattern raises two general questions: 1) Do a few large-effect genes contribute to latitudinal variation in a trait? 2) Is the thermal plasticity of different traits regulated pleiotropically? To address the questions, we crossed individuals of Plantago lanceolata derived from northern and southern European populations. Individuals naturally exhibited high and low thermal plasticity in floral reflectance and flowering time. We grew parents and offspring in controlled cool- and warm-temperature environments, mimicking what plants would encounter in nature. We obtained genetic markers via genotype-by-sequencing (ddRADseq), produced the first recombination map for this ecologically-important non-model species, and performed QTL mapping of thermal plasticity and single-environment values for both traits. We identified a large-effect QTL that largely explained the reflectance plasticity differences between northern and southern populations. We identified multiple smaller-effect QTLs affecting aspects of flowering time, one of which affected flowering time plasticity. Results indicate that the genetic architecture of thermal plasticity in flowering is more complex than for reflectance. One flowering time QTL showed strong cytonuclear interactions under cool temperatures. Reflectance and flowering plasticity QTLs did not colocalize, suggesting little pleiotropic genetic control and freedom for independent trait evolution. Such genetic information about the architecture of plasticity is environmentally important because it informs us about the potential for plasticity to offset negative effects of climate change.

Floral reflectance was measured as the average of % light reflected at 850nm for two spectral scans conducted on a single pre-flowering spike using a spectrophotometer with an integrating sphere.

Flowering time was the number of days from induction (day 168) to the complete emergence of the first flowering spike.

Readme.txt

Data: REF_FT_Phenotypic_Data.xlsx
- Phenotypic data for all individuals as summarized in Table 1 organized into 5 tabs.
- Tab 'Parents.REF' - Reflectance Data from F0 Parent clones, and mean values.  Columns 'Genotype' - A (Italy), B (Denmark), C (France), J (Sweden); 'Clone' - Clone Number or MEAN; 'Temperature' - Cool, Warm, or Mean; 'Cool' - Reflectance value in Cool; 'Warm' - Reflectance value in Warm; 'Plasticity' - Plasticity Value
- Tab 'F1.REF' - Reflectance Data from F1 genotypes. Columns 'Genotype'- BA and CJ are F1-parents; 'Family' - Parents listed as female x male, as in Figure 2; 'Generation' - F1; 'Temperature' - Cool, Warm; 'F1_ref' - Single environment genotypic mean calculated as the average of clonal replicates; 'Plast_REF' - Mean plasticity calculated as mean warm value - mean cool value
- Tab 'F2.REF' - Reflectance Data from F2 genotypes. Columns 'Genotype'- Individual; 'Cytoplasm' - N or S; 'Generation'- F2; 'Cool Reflectance' - Cool environment genotypic mean calculated as the average of clonal replicates; 'Warm Reflectance' - Warm environment genotypic mean calculated as the average of clonal replicates; 'PlasticityF2' - Mean plasticity calculated as mean warm value - mean cool value
- Tab 'Parents.FT' - Flowering Time Data from F0 Parent clones, and mean values.  Columns 'Genotype' - A (Italy), B (Denmark), C (France), J (Sweden); 'Clone' - Clone Number or MEAN; 'Temperature' - Cool, Warm, or Mean; 'Cool' - Flowering Time value in Cool; 'Warm' - Flowering Time value in Warm; 'Plasticity' - Plasticity Value
- Tab 'F1_F2.FT' - Flowering Time Data from F1 and F2 genotypes. Columns 'Genotype'- BA and CJ are F1-parents; 'Family (for F1s) Cytoplasm (for F2s)' - Parents listed as female x male for F1s, Cytoplasm N or S for F2s (See Fig 2); 'Generation'- F1 or F2; 'Temperature' - Cool, Warm; 'F1_FT'- Flowering Time genotypic mean calculated as the average of clonal replicates of F1s; 'F2_FT'- Flowering Time genotypic mean calculated as the average of clonal replicates of F2s; 'Plast_FT' - Mean plasticity calculated as mean warm value - mean cool value 

Script: PVE.R - R Script for performing PVE calculations
Data: PVE_Data.CSV - Raw data of genotypic means for parent and F2 genotypes.
- Columns: 'Genotype' - Genotype of measurement, 'Generation' - South or North for F0 parents, F2 for F2 genotypes
- Columns: Data columns indicate trait and environment (or plasticity) as Trait.Environment; 'Reflectance.Cold', 'Reflectance.Warm', 'Reflectance.Plasticity, 'FT.Cold', 'FT.Warm', 'FT.Plasticity'
- Values indicate mean trait value. For F0 genotypes mean trait value represents mean of a single clone.  For F2 genotypes mean trait value represents mean of all clonal values.  See methods for details.
 
Script: Recombination_Mapping.R
Data: Recombination_Mapping.txt - Raw data of 323 genetic markers for 104 F2 individuals with highest sequencing coverage. File structure is as required by OneMap 2.0-4 package that runs in R.

Script: qtl_FT.R
Script: qtl_Reflectance.R
Data: ALL_DATA.qtl.csv; File structure is as required by qtl package that runs in R. 
- Columns: 'Genotype'- F2 Genotype, 'Cytoplasm' - N or S cytoplasm, 
- Columns: Data columns indicate trait and environment (or plasticity) as Trait.Environment; 'Reflectance.Cold', 'Reflectance.Warm', 'Reflectance.Plasticity, 'FT.Cold', 'FT.Warm', 'FT.Plasticity'
- Columns: '393', '99', '493', etc.: These are the DNA markers from the recombination map (As in Figure 5). 
Data: ALL_DATA.qtl_N.csv; Same as ALL_DATA.qtl.csv except F2s with Southern cytoplasm (S) are excluded. 
Data: ALL_DATA.qtl_S.csv; Same as ALL_DATA.qtl.csv except F2s with Northern cytoplasm (N) are excluded.