Field experiment and phenotyping
The details of the creation of this population were described in Milano et al. (2016). Briefly, the grandparents of the mapping population were derived from highly divergent southern lowland and northern upland ecotypes. The population was developed by initial crosses between AP13 (A) x DAC6 (B) and WBC3 (C) x VS16 (D). AP13 and WBC3 are genotypes clonally derived from an individual selected from the lowland cultivar ‘Alamo’ (southern Texas accession) and an individual from naturally occurring population ‘West Bee Cave’ (central Texas accession), respectively. DAC6 and VS16 are genotypes clonally derived from individuals selected from the upland cultivars ‘Dacotah’ and ‘Summer’ (both northern upland accessions), respectively. The F1hybrids of each of those crosses were then intercrossed reciprocally to produce the four-way outbred mapping population.
The grandparents, F1 hybrid parents, and the F2 progeny were propagated by dividing plants manually to produce 10 clones, each of which was maintained in a 3.8-L pot at the Brackenridge Field Laboratory, Austin, TX in 2013-2015. One replicate of each of the mapping progeny genotypes, along with multiple replicates of grandparents and F1 parents, were transplanted from May to July of 2015 at 10 field sites. In three of the 10 sites, 370 extra genotypes were planted. The 10 field sites cover 17 degrees of latitude from South Texas to South Dakota (Figure 1A). Detailed information of the 10 field sites, including latitude, longitude and soil type is provided in Table 1. The annual mean temperature at the 10 sites in 2016 ranged from 10.4 °C in the north to 20.7 °C in the south, and the total rainfall varied from 574 mm to 1440 mm (Figure 1B, data are from local weather station or from NOAA if local weather data are not available; the weather station or NOAA link is included in Table 1). To control weeds, each field site was covered with one layer of weed barrier cloth (Dewitt, Sikeston, MO). Holes were cut into the weed cloth in a honeycomb fashion. Plants were randomized into the holes, with each plant having four nearest neighbors each located 1.56m away from each other. A row of border plants was planted at every edge position of the field to minimize edge effects. The border plants were derived from rhizome plugs obtained from an approximately 10-year-old stand of Alamo switchgrass. Plants were well watered in the field during the summer of 2015 to facilitate establishment and all phenotypes were collected in 2016.
Three panicles were cut from each plant at full maturity. Panicle length (PL in mm), primary branching number (PBN), and secondary branching number (SBN) were assessed at the end of the growing season. A diagram depicting these phenotypes is presented in Figure 2, with representative images of panicles from the four grandparents. PL was measured on the primary panicle from the base of the first primary branch to the top of the panicle. PBN was counted as the total number of branches along the primary rachis. Due to the numerous secondary branches in switchgrass, SBN in our study referred to the total number of secondary branches on the lowest primary branch of the panicle (Figure 2). In total, over 10,000 separate panicle morphology measurements were collected in our study. The phenotypic data (i.e., average values) for each genotype at each field site are provided in Supplemental Table S1.