Accession number Species Taxonomic. Reproductive mode. group Altitude (m) Latitude Longitude

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1 Supplementary Table S1. number, species, and taxonomic group (as per Renvoize et al., 1996) of each of the 261 genotypes included in the study. Geographical information about the collection site, reproductive mode, and other identifiers for the accession are also provided where available B. ruziziensis SX R135, BRA B. ruziziensis unknown unknown nd BRA B. ruziziensis SX ILCA B. ruziziensis SX R101, BRA , ILCA B. ruziziensis SX R102, BRA , ILCA B. ruziziensis SX R108, BRA , ILCA B. ruziziensis SX B. ruziziensis SX R105, BRA , ILCA B. ruziziensis SX R109, BRA B. ruziziensis SX B B. ruziziensis SX D024, BRA , ILCA B. ruziziensis SX ILCA B. ruziziensis SX R112, BRA , ILCA BRX B. ruziziensis 5 na # na na SX MOK8-4 B. ruziziensis 5 na na na SX MOK8-6 B. ruziziensis 5 na na na SX MOK8-7 B. ruziziensis 5 na na na SX 606 B. decumbens 5 unknown unknown unknown AP cv. Basilisk, D062, BRA , ILCA B. decumbens AP D067, BRA B. decumbens AP D002, BRA , ILCA B. decumbens SX D006, BRA , ILCA B. decumbens AP D007, BRA , ILCA B. decumbens AP D011, BRA , ILCA B. decumbens AP BRA , ILCA B. decumbens AP D015, BRA , ILCA B. decumbens AP ILCA B. decumbens SX D017, BRA , ILCA B. decumbens SX D029, BRA , ILCA B. decumbens SX D035, BRA , ILCA B. brizantha 5 unknown unknown unknown nd BRA B. brizantha 5 unknown unknown unknown AP cv. Marandu, B030, BRA B. brizantha AP BRA B. brizantha AP BRA

2 6684 B. brizantha AP B038, BRA B. brizantha AP B040, BRA B. brizantha AP B033, BRA B. brizantha AP B034, BRA B. brizantha AP B039, BRA B. brizantha nd 6780 B. brizantha 5 unknown unknown unknown nd ILCA B. brizantha AP B102, BRA , ILCA B. brizantha AP B103, BRA , ILCA B. brizantha AP B185, BRA , ILCA B. brizantha AP B109, BRA , ILCA B. brizantha AP B111, BRA , ILCA B. brizantha nd ILCA B. brizantha AP B186, BRA , ILCA B. brizantha AP B213, BRA , ILCA B. brizantha nd ILCA B. brizantha AP B121, BRA , ILCA B. brizantha nd ILCA B. brizantha AP B122, BRA B. brizantha AP B187, BRA , ILCA B. brizantha AP B219, BRA , ILCA B. brizantha nd BRA , ILCA B. brizantha AP B057, BRA , ILCA B. brizantha AP B128, BRA , ILCA B. brizantha AP B059, BRA , ILCA B. brizantha AP B221, BRA , ILCA B. brizantha AP B060, BRA , ILCA B. brizantha nd ILCA B. brizantha AP B134, BRA , ILCA B. brizantha AP B135, BRA , ILCA B. brizantha AP B136, BRA , ILCA B. brizantha nd BRA , ILCA B. brizantha AP B064, BRA , ILCA B. brizantha AP B067, BRA , ILCA B. brizantha AP B138, BRA , ILCA-13610

3 16309 B. brizantha AP B140, BRA , ILCA B. brizantha AP B142, BRA , ILCA B. brizantha AP B188, BRA , ILCA B. brizantha AP B224, ILCA B. brizantha AP B150, BRA , ILCA B. brizantha AP B076, BRA , ILCA B. brizantha AP B078, BRA , ILCA B. brizantha AP B080, BRA , ILCA B. brizantha AP B082, BRA , ILCA B. brizantha AP B084, BRA , ILCA B. brizantha AP B085, BRA , ILCA B. brizantha AP B232, BRA , ILCA B. brizantha AP B235, BRA , ILCA B. brizantha nd BRA , ILCA B. brizantha AP B168, BRA , ILCA B. brizantha nd ILCA B. brizantha AP B192, BRA , ILCA B. brizantha nd BRA , ILCA B. brizantha AP B171, BRA , ILCA B. brizantha AP H118, BRA B. brizantha AP B173, BRA B. brizantha nd B. brizantha AP B B. brizantha nd B. brizantha nd B. brizantha AP B175, BRA B. brizantha nd ILCA B. brizantha AP B209, BRA B. brizantha nd ILCA B. brizantha AP BRA B. brizantha nd BRA B. brizantha AP B255, ILCA B. brizantha AP B097, BRA B. brizantha AP B098, BRA B. brizantha AP B256, ILCA-16182

4 16835 B. brizantha nd BRA B. brizantha nd ILCA B. brizantha nd BRA B. brizantha AP B210, BRA B. brizantha AP B258, BRA B. brizantha nd B. brizantha nd B. brizantha AP B259, ILCA B. brizantha AP B208, BRA , ILCA B. brizantha AP B B. brizantha nd B. brizantha nd B. brizantha nd B. brizantha nd BRA , ILCA B. brizantha AP B200, BRA , ILCA B. brizantha AP ILCA B. brizantha AP B. brizantha AP B. brizantha AP ILCA B. brizantha AP B260, ILCA B. brizantha AP ILCA B. brizantha nd ILCA B. brizantha AP ILCA B. brizantha AP B B. brizantha AP B. brizantha AP B B. brizantha AP B296, ILCA B. brizantha AP B293, BRA , ILCA B. brizantha AP B182, BRA , ILCA B. brizantha AP B183, BRA , ILCA B. brizantha AP BRA , ILCA B. brizantha AP B266, BRA , ILCA B. brizantha AP B267, ILCA B. brizantha AP B268, ILCA B. brizantha AP ILCA-00767

5 26393 B. brizantha AP B271, ILCA B. brizantha nd BRA , ILCA B. brizantha AP B275 SX14/0058 Interspecific Brachiaria hybrid 5 na na na SX SX14/0129 Interspecific Brachiaria hybrid 5 na na na SX SX14/0184 Interspecific Brachiaria hybrid 5 na na na SX SX14/0253 Interspecific Brachiaria hybrid 5 na na na SX SX14/0307 Interspecific Brachiaria hybrid 5 na na na SX SX14/0518 Interspecific Brachiaria hybrid 5 na na na SX SX14/0576 Interspecific Brachiaria hybrid 5 na na na SX SX14/0775 Interspecific Brachiaria hybrid 5 na na na SX SX14/0892 Interspecific Brachiaria hybrid 5 na na na SX SX14/1071 Interspecific Brachiaria hybrid 5 na na na SX Interspecific Brachiaria hybrid 5 na na na AP cv. Mulato Interspecific Brachiaria hybrid 5 na na na AP cv. Mulato II BR02/1752 Interspecific Brachiaria hybrid 5 na na na AP cv. Cayman BR02/1794 Interspecific Brachiaria hybrid 5 na na na AP cv. Cobra JIRCAS 185 Interspecific Brachiaria hybrid 5 na na na AP JIRCAS 203 Interspecific Brachiaria hybrid 5 na na na AP JIRCAS 226 Interspecific Brachiaria hybrid 5 na na na AP B. dura nd 675 B. humidicola 6 unknown unknown unknown AP BRA B. humidicola 6 unknown unknown unknown AP cv. Tully, cv. Humidicola, BRA , 6133 B. humidicola AP cv. Llanero, H125, BRA , ILCA B. humidicola 6 unknown unknown unknown nd BRA , ILCA B. humidicola AP BRA B. humidicola nd BRA , ILCA B. humidicola AP H004, BRA , ILCA B. humidicola AP H005, BRA , ILCA B. humidicola AP H020, BRA , ILCA B. humidicola AP H019, BRA , ILCA B. humidicola AP H042, BRA B. humidicola AP H006, BRA B. humidicola nd BRA B. humidicola AP H007, BRA

6 16871 B. humidicola AP H008, BRA B. humidicola AP B. humidicola AP H022, BRA B. humidicola AP BRA B. humidicola AP H023, BRA B. humidicola AP H024, BRA B. humidicola AP H009, BRA B. humidicola AP H010, BRA B. humidicola AP H012, BRA B. humidicola AP H025, BRA B. humidicola AP H044, BRA B. humidicola AP H045, BRA B. humidicola AP H013, BRA B. humidicola AP H026, BRA B. humidicola AP H027, BRA B. humidicola AP H014, BRA B. humidicola AP H028, BRA B. humidicola AP B. humidicola AP H029, BRA B. humidicola nd BRA B. humidicola AP H046, BRA B. humidicola SX H031, BRA , ILCA B. humidicola AP cv. Tupi, H016, BRA , ILCA B. humidicola AP B. humidicola AP B. humidicola AP H017, BRA B. humidicola AP H018, BRA , ILCA B. humidicola 6 unknown unknown unknown AP H047, BRA , ILCA B. humidicola AP H120, BRA , ILCA B. humidicola AP BRA , ILCA B. humidicola nd BRA , ILCA B. humidicola AP H122, BRA , ILCA B. humidicola AP H033, BRA , ILCA B. humidicola AP H034, BRA , ILCA B. humidicola AP H035, BRA , ILCA-00734

7 26414 B. humidicola AP H036, BRA , ILCA B. humidicola AP H037, BRA , ILCA B. humidicola AP H038, BRA , ILCA B. humidicola AP BRA , ILCA B. humidicola AP H039, BRA , ILCA B. humidicola AP H040, BRA , ILCA B. humidicola nd BRA , ILCA B. humidicola 6 unknown unknown unknown nd BRA B. humidicola AP BRA , ILCA BhSX12/001 B. humidicola hybrid 6 na na na SX BhSX12/025 B. humidicola hybrid 6 na na na SX BhSX12/062 B. humidicola hybrid 6 na na na SX BhSX12/121 B. humidicola hybrid 6 na na na SX BhSX12/158 B. humidicola hybrid 6 na na na SX BhSX12/214 B. humidicola hybrid 6 na na na SX BhSX12/261 B. humidicola hybrid 6 na na na SX BhSX12/328 B. humidicola hybrid 6 na na na SX BhSX12/375 B. humidicola hybrid 6 na na na SX BhSX12/474 B. humidicola hybrid 6 na na na SX B. bovonei nd BRA B. bovonei nd ILCA B. dictyoneura nd ILCA B. dictyoneura nd ILCA B. jubata nd BRA B. jubata nd BRA , ILCA B. jubata nd BRA , ILCA B. jubata nd BRA , ILCA B. jubata nd BRA , ILCA B. jubata nd BRA , ILCA B. jubata nd BRA , ILCA B. jubata nd BRA , ILCA B. jubata nd ILCA B. platynota nd ILCA B. platynota nd BRA , ILCA B. subulifolia nd BRA

8 16966 B. subulifolia nd BRA B. nigropedata nd B. nigropedata nd ILCA B. nigropedata nd ILCA B. nigropedata nd ILCA B. nigropedata nd B. nigropedata nd ILCA B. nigropedata nd B. nigropedata nd B. serrata nd B. serrata nd BRA B. arrecta nd BRA B. arrecta nd BRA B. eruciformis nd cv. Paikaji P. maximum na na na na nd cv. Umak P. maximum na na na na nd cv. Massai P. maximum na na na na nd cv. Mombaça P. maximum na na na na nd CIAT Genetic Resources Program ( accession number for each genotype (3-, 4-, or 5-digit number). CIAT breeding code (BRX 44-02, SX14, BhSX, BR02, or BhSX12); JIRCAS identifier (JIRCAS or MOK prefix), or cultivar name are given for those genotypes not available in the CIAT genebank. Species of each accession according to the CIAT Genetic Resources Program database ( Interspecific Brachiaria hybrids were developed from crosses between the agamic complex species (B. ruziziensis, B. decumbens, B. brizantha ) Taxonomic groups, according to Renvoize et al. (1996) AP = apomict, SX = sexual, nd = not determined. mode was determined with primer pair p779/p780, a diagnostic marker for apomixis in Brachiaria, following Worthington et al. (2016). # na = not applicable.

9 Supplementary Table S2. Primer sequences, number of polypmorphic bands used in the study, range of polymorphic band sizes, repeat motifs, melting temperatures, and polymorphism information content (PIC) for each of the 39 simple sequences repeat (SSR) markers. Number of Tm values ( C) SSR primer combination polymorphic bands Band size (bp) Forward primer sequence Reverse primer sequence Repeat motif Forward primer Reverse primer PIC BD GAGGAAGGCAGGAGGAAGTT TGCATGCATGACAACAAAGA (AG)^ BD TTGGTTCTGAGAAGGTTGGTG TGGAAACTTGGGAGGATCTG (AG)^ BD GCGATTCACAAGCCAGTCTC GCAGAGCCAGGATCAGAAAC (AT)^ BD TGGAAGAGGGAACTGTGACC TGAGGTTTCGGCTAGACCAG (GT)^ BD GAACGGTGCAAGTGGAAGAT CAGCTGCAACAAGTATGTTCG (AT)^ BD ATCGTGCTCACCTTCTTCGT CTAGGCGGCTCATCGTAAAC (CG)^ BD GCTGATCAGGCGTTTAGCTC CGTAGCCCAACTTTGAGAGG (CT)^ BD TGAAGCTGGTGCACTCTTTG GGAGTGTATCCTTCAGCAACC (CT)^ BD AAGGACTTTGCGTGCACTTT AGGGCACTGCACTGAAGACT (CT)^ BD ATTTTGTGCCTCCCTACGTG GGACGAAGAACATACGCATTC (GT)^ BD GCTGCATAGATGCATTCACAA CTGCTGCTGTGGTACATGCT (AT)^ BD CACAGGGACTCACTAGGTTGC AATTGCATGCTTTCCTCCAG (GT)^ BD GCGAAACAACACCATGTCAC GCTCGGCTACAACTTCCTTTT (GT)^ BD GCAATAGCCCAAGCTGCTAC GCAATAGCACCAAGACACAGTC (CT)^ BH CGCTATGAACTTCTGCACCA AAGGCCTCAATCTGCACAAC (ACTCCG)^ BH TTTTGCACTTCCTCCAGCTT CTTATGGCTGTCGAGCACAA (GCTTTT)^ BH GGCCTCAAGCTCTGCTATTC GCACGACTCCAACAACAAGA (CGGAGT)^ BTe CCAGTGCCTCTACAGGACCA GCTCAAGCTCAGGGATGAAG (ATCC)^ BTr AGGACGAGGAGGAAGAGGAG AATGCGCTTGCGAATAAAGT (AAG)^ BTr CAAAGGGTGAAGCCAAGGTA TCATGAGTGGTCTTTAGCTCCTC (GAT)^ BTr GGTGGTGGTAGCTTTTCCAG TACACCAGCAACCAAACCAA (GCT)^ BTr CTCATCGGCGTCCAGATATT CACATGCCTTCAACCAGGA (CCT)^ BTr TTTCTTTGGTGTGCAGGTTG GCGATCTCCTTGAGGTCGTA (AGC)^ BTr ACATGCAAGCAGCAGCATAG GGTTCATGCCCATCTTGATT (ATC)^ BTr ACCGAGCACGGTGAACTTAG AATGAGGTGGCTCATCTTGTG (GAT)^ BTr GCATGCATTGCTGTTGTTCT TGCGTATGCTTCGTGTCTTC (ACC)^ BTr GTTGATGAACGTCCTGAGCA CTGAGAATTGGTCCGGAGAG (CGG)^ BTr CCGGCAGGTACTACATGGTC GTGGAGGCTTGTAAGGTTGC (AAG)^ BTr AGGAAGTACTCGCCGTGGT CTTCATTGAGCGTCACTTGC (GCT)^ BTr AACAAGCGCACGAACCATAC CAGCAAGAGGACCAGAGGAG (ACT)^ BTr TACAACCCTGCTGCTGATGT GGAGTCTCCTCCGTTTCCTT (AAG)^ BTr GTCGTACATACCGCCTCCAT GCGCCAAAGAGAAATGAAGA (ACC)^ BTr GATGAACAGCCGCTAGGAAA CGTGAGATCTTTTTCCCTCCT (AGC)^ BTr GCCCGTAGAAATCGTGAAAA GCCTTCCATTTCCAAGCTAC (GAT)^ BTr ACCGACCTTGCTGCTTATGT GTCCAAGGAATGTGGCCTAA (ACT)^

10 SSR primer combination Number of polymorphic bands Band size (bp) Forward primer sequence Reverse primer sequence Repeat motif Forward primer Reverse primer PIC BTr GTCTCGCCAATAACCACGTT AGCCTGAAAATGGTGCAAAG (CTT)^ BTr GCACGAAGCTAGCGAAGAG CGTCCTCACCGCTCAAGT (GGT)^ BTr CGAGGGTGCTCTTGATCTTC GCGTCTCTTACCTTGCTTGC (CCG)^ BTr AGGATTTTGGCTCTGGGTTT CGTAGTAAAGAAGTGAAATGAG (ATC)^

11 Supplementary Table S3. Average number of marker bands present for each of the 14 Brachiaria species evaluated as well as the interspecific Brachiaria hybrid genotypes and the Panicum maximum outgroup Species Taxonomic group Average number of marker bands present B. ruziziensis 5 36 B. decumbens 5 42 B. brizantha 5 51 Interspecific Brachiaria hybrid 5 47 B. dura 5 30 B. humidicola 6 39 B. bovonei 6 35 B. dictyoneura 6 34 B. jubata 6 34 B. platynota 6 44 B. subulifolia 6 31 B. nigropedata 2 30 B. serrata 2 48 B. arrecta 3 26 B. eruciformis 7 49 P. maximum na 32 Species of each accession according to the CIAT Genetic Resources Program database ( Interspecific Brachiaria hybrids were developed from crosses between the agamic complex species (B. ruziziensis, B. decumbens, B. brizantha ) Taxonomic groups, according to Renvoize et al. (1996)

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