Supplement 2. PCR Protocols for the Ampliconsused in the Study

Supplementary materials

Supplement 1. List of the lines from the association panel. For each line is given the origin, the name of the line, the sum of temperature for flowering time in degree-days and the maturity group that was used to divide the panel into three groups for per se trials.

Supplement 2. PCR protocols for the ampliconsused in the study

a.The PCR reactions for the two amplicons containing respectively the mite and CGindel587 were performed in 15L volumes. For the first amplicon, the volumes contained 9.5L of bidistilled water, 2L of Taq polymerase 10X (Qiagen), 0.4L of MgCl2 (25mM), 0.4L of each dNTP (10mM), 0.1L of each forward and reverse primer (100M), 2.5L of Taq M (0.5U/L) and 2L of genomic DNA. The PCR program was 4min at 94°C, followed by 11 cycles of 1min at 94°C, 1 min from 64 to 54°C and 1min30s at 72°C, followed by 25 cycles of 1min at 94°C, 1min at 54°C and 1min30s at 72°C, followed by 4min at 72°C and 10min at 4°C. For the second amplicon, the volumes contained 9.08L of bidistilled water, 2L of Taq polymerase 10X (Qiagen), 1.2L of MgCl2 (25mM), 0.4L of each dNTP (10mM), 0.16L of each forward and reverse primer (100M), 2L of Taq M (0.5U/L) and 2L of genomic DNA. The PCR program was 4min at 94°c, followed by 11 cycles of 30s at 94°c, 30s at 66 to 56°C and 1min30s at 72°C, followed by 25 cycles of 30s at 94°C, 30s at 56°C and 1min30s at 72°C, followed by 5min at 72°C and 5min at 4°C.

b.. The PCR reactions for the 101 amplicons developed over the 13 regions of the genome and the other 10 in the Vgt1 region were performed in 15L volumes for the mini-panel experiments and 10L for the whole panel experiments. The volumes contained 1X PCR buffer (invitrogen, Carlsbad, CA), 0.6U of Taq polymerase (Invitrogen, Platinum), 1.5mM of MgCl2, 0.2mM of each dNTP, 0.2mM of each forward and reverse primers and 5L of genomic DNA. The PCR program was 2min at 94°c, followed by 10 cycles of 30s at 94°c, 30s at 62°c and 2min at 72°c, followed by 25 cycles of 30s at 94°c, 30s at 57°c and 2min at 72°c, followed by 5min at 72°c.

Supplement 4. r2for marker distant from 0 to 1000bp, for the whole panel.A total of 1128 polymorphisms were used on the graphs; (a) is based on all polymorphisms (all allelic frequencies), (b) is based on markers with MAF>0.2 and (c) is based on marker pairs for which at least one marker had a MAF<0.2.

Supplement 6.Comparison of the different models for association mapping for hybrids FFLW. Simple=Generalized Linear Model; Q3=STRUCTURE 3 groups; PCA10=Eigenstrat 10 axis; Kemma= Mixed linear model with Kemma; KLoiselle=Mixed Linear Model with KLoiselle