Amplification Results: 1, Band of the Expected Size; 0, Absence of Amplification Products

Supplementary Fig. 1. Alignment of GenBank 16S rRNA gene sequences of members of Actinomycetes taxa used to evaluate the specificity of the Actinopolyspora-specific primers AcpF/AcpR. Shaded boxes indicate conserved positions. Shaded boxes indicate conserved positions. 5’ 261. .284 3’ 5’ 447 . .469 3’

AcpF CGGTGAGGGCGTACCAAGGCGATG AcpR AGCCCCTACCGTCACA------TTCGTCA Actinopolyspora mortivallis DSM 44261T (DQ883812) TGGTGCGGTGAGGGCGTACCAAGGCGATGACGG TTAGCCCCTACCGTCACA------TTCGTCAGGGCCG Actinopolyspora halophila ATCC 27976T (X54287) TGGTGCGGTGAGGGCGTACCAAGGCGATGACGG TTAGCCCCTACCGTCACA------TTCGTCAAGGGCC Kutzneria kofuensis NRRL B-24061T (AF114801) TGGTGGGGTAGTGGCCTACCAAGGCGACGACGG TCTGCAGGTACCGTCACT------T-GCGCTTCGTCCCTGCTG Saccharothrix australiensis NRRL 11239T (AF114803) TGGTGGGGTGATGGCCTACCAAGGCGACGACGG TCTGCAGGTACCGTCACT------C-ACGCTTCGTCCCTGCTG Crossiella equi NRRL B-24104T (AF245017) TGGTGGGGTAATGGCCTACCAAGGCGACGACGG TCTGTACCTACCGTCACT------T-TCGCTTCGTCGGCACTG Streptoalloteichus hindustanus IFO15115 (D85497) TGGTGGGGTAGTGGCCTACCAAGGCGACGACGG TCTGCAGGTACCGTCACT------NTGCGCTTCGTCCCTGCTG Actinoalloteichus cyanogriseus IFO14455 (AB006178) TGGTGGGGTAATGGCCTACCAAGGCGACGACGG TCTGCGCCTACCGTCAC------TTTCGCTTCTTCGGCGCTG Pseudonocardia asaccharolytica DSM 44247T (Y08536) TGGTGGGGTGATGGCCCACCAAGGCGACGACGG TCTCCCCCTACCGTCACCCC------CAAAAGGGCTTCGTCGGAAGCG Pseudonocardia kongjuensis LM 157T (AJ252833) TGGTGGGGTGATGGCCTACCAAGGCGGTGACGG TCTCCAGGTACCGTCAC------AAAAGCTTCGTCCCTGGCG Kibdelosporangium aridum subsp. largum DSM 44150T (AJ512463) TGGTGGGGTGATGGCCTACCAAGGCGACGACGG TCTCCAGGTACCGTCAC------TCTCGCTTCGTCCCTGGCG Nonomuraea africana DSM 43748T (AJ269555) TGGTGGGGTAGTGGCCTACCAAGCCGACGACGG TCTGCAGGTA----CACGTC------AACTTCGTCCCTGCTG Thermopolyspora flexuosa DSM 43186T (AY039253) TGGTGGGGTAACGGCCTACCAAGGCGACGACGG TCCGCAGGTA----CACGTC------ACCTTCGTCCCTGCCG Thermobispora bispora ATCC 19993 (U58523) TGGTGGGGTGACGGCCTACCAAGGCGATGACGG TCTGCAGGTA----CACGTC------AACTTCGTCCCTGCTG Actinomadura atramentaria DSM 43919T (AJ420138) TGGTGGGGTGATGGCCTACCAAGGCGACGACGG TCTGCAGGTACCGTCACGTT------AGCTTCGTCCCTGCTG Nocardiopsis alba DSM 43377T (X97883) TGGTGGGGTAACGGCCTACCAAGGCGATTACGG TCCCCACCTACCGTCAACCCCGAGAGAACCCGGAGCCTGCGTTGGTGGTA Streptomonospora alba YIM90003T (AF462347) TGGTGGGGTGAAGGCCTACCAAGGCGATTACGG TCCCCACCTACCGTCAACCCGCACAGAAGCGCGGGCCTGCGTCGGTGGTA Micromonospora auratinigra TT1-11T (AB159779) TGGTGGGGTGATGGCCTACCAAGGCGACGACGG TCTGCAGGTACCGTCACTTT------CGCTTCGTCCCTGCTG Actinoplanes consettensis JCM 7624T (AB048214) TGGTGGGGTAATGGCCTACCAAGGCGAGGACGG TCTGCAGGTACCGTCACTTT------CGCTTCGTCCCTGCTG Kribbella flavida KACC 20248T (AY253863) TGGTGGGGTAATGGCCTACCAAGGCGTCGACGG TCTGCAGGTACCGTCACTCT------CGCTTCGTCCCTGCTG Microlunatus phosphovorus NM-1T (D26169) TGGTGAGGTAACGGCTCACCAAGGCTTCGACGG TCTGCTCCTACCGTCACTTG------CGCTTCGTCGGAGCTG Jiangella gansuensis YIM 002T (AY631071) TGGTGGGGTAGTGGCCTACCAAGGCGATGACGG TCTGCGCCTACCGTCACCCG------AAGGCTTCGTCAGCGCTG Streptomyces albus subsp. albus DSM 40313T (AJ621602) TGGTGGGGTGATGGCCTACCAAGGCGACGACGG TCTGCAGGTACCGTCACTC------GCGC--TTCTTCCCTGCTG Amycolatopsis balhimycina DSM 44591T (AJ508239) TGGTGGGGTAGTGGCCTACCAAGGCGACGACGG TATCCAGGTACCGTCACTT------GCGC--TTCGTCCCTGGCG

Amycolatopsis rubida 13.4T (AF222022) TGGTGGGGTAGTGGCCTACCAAGGCGACNACGG TATCCAGGTACCGTCACTC------TCGC--TTCGTCCCTGGCG

Prauserella halophila YIM 90001T (AF466190) TGGTGGGGTGATGGCCTACCAAGGCGGTGACGG TCTCCAGGTACCGTCAGTA------TCACCCTTCGTCCCTGGCG

Prauserella alba YIM 90005T (AF435077) TGGTGGGGTGATGGCCTACCAAGGCGGTGACGG TCTCCAGGTACCGTCAGTA------TCACCCTTCGTCCCTGGCG

Saccharomonospora xinjiangensi DSM 44391T (AJ306300) TGGTGGGGTGATGGCCTACCAAGGCGACGACGG TCTCCCGGTACCGTCACTT------TCGC--TTCGTCCCGGGCG Thermocrispum municipale DSM 44069 (X79184) TGGTGGGGTGATGGCCTACCAAGGCGGTGACGG TCTACCCCTACCGTCACTC------TCGC--TTCGTCGGGGTCG Saccharopolyspora flava AS4.1520T (AF154128) TGGTGGGGTAGTGGCCTACCAAGGCGACGACGG TCTACAGGTACCGTCAGTT------GCCC--TTCGTCCCTGTCG

Supplementary Fig. 2. Alignment of GenBank 16S rRNA gene partial sequences of new wild type isolates used in multiplex-PCR experiments to evaluate the specificity of the Actinopolyspora-specific primers AcpF/AcpR. Shaded boxes indicate conserved positions. Shaded boxes indicate conserved positions.

5’ 261. .284 3’ 5’ 447 . .469 3’

AcpF CGGTGAGGGCGTACCAAGGCGATG AcpR AGCCCCTACCGTCACA------TTCGTCA YIM 90588 GGTGCGGTGAGGGCGTACCAAGGCGATGACG CTTTAGCCCCTACCGTCACA------TTCGTCAGGGCC YIM 90589 GGTGCGGTGAGGGCGTACCAAGGCGATGACG CTTTAGCCCCTACCGTCACA------TTCGTCAGGGCC YIM 90601 GGTGCGGTGAGGGCGTACCAAGGCGATGACG CTTTAGCCCCTACCGTCACA------TTCGTCAGGGCC YIM 90617 GGTGCGGTGAGGGCGTACCAAGGCGATGACG CTTTAGCCCCTACCGTCACA------TTCGTCAGGGCC YIM 90600 GGTGCGGTGAGGGCGTACCAAGGCGATGACG CTTTAGCCCCTACCGTCACA------TTCGTCAGGGCC YIM 90570 GGTGCGGTGAGGGCGTACCAAGGCGATGACG CTTTAGCCCCTACCGTCACA------TTCGTCAGGGCC YIM 90479 GGTGCGGTGAGGGCGTACCAAGGCGATGACG CTTTAGCCCCTACCGTCACA------TTCGTCAGGGCC YIM 90485 GGTGCGGTGAGGGCGTACCAAGGCGATGACG CTTTAGCCCCTACCGTCACA------TTCGTCAGGGCC YIM 90829 GGTGCGGTGAGGGCGTACCAAGGCGATGACG CTTTAGCCCCTACCGTCACA------TTCGTCAGGGCC

YIM 90567 GGTGGGGTAACGGCCTACCAAGGCGATTACG ATTCCCCACCTACCGTCAACCCGCACAGAAGCACGGGCCTGCGTCGGTGGT

YIM 90568 GGTGGGGTAACGGCCTACCAAGGCGATTACG ATTCCCCACCTACCGTCAACCCGCACAGAAGCACGGGCCTGCGTCGGTGGT

YIM 90571 GGTGGGGTAACGGCCTACCAAGGCGATTACG ATTCCCCACCTACCGTCAACCCGCGCAGAAACGCGGGCCTGCGTCGGTGGT

YIM 90632 GGTGGGGTGAAGGCCTACCAAGGCGATTACG ATTCCCCACCTACCGTCAACCCGCACAGAAGCGCGGGCCTGCGTCGGTGGT

YIM 90554 GGTGGGGTGAAGGCCTACCAAGGCGATTACG ATTCCCCACCTACCGTCAACCCGCGCAGAAACGCGGGCCTGCGTCGGTGGT

YIM 90487 GGTGGGGTGAAGGCCTACCAAGGCGATTACG ATTCCCCACCTACCGTCAACCCCGGGAGAACCCGGGGCCTGCGTGAGTGGT

YIM 90494 GGTGGGGTGAAGGCCTACCAAGGCGATTACG ATTCCCCACCTACCGTCAACCCCCGGAGAACCGGGAGCCTGCGTTGGTGGT

YIM 90562 GGTGGGGTGAAGGCCTACCAAGGCGATTACG ATTCCCCACCTACCGTCAACCCCCAGAGAACTGGGAGCCTGCGTTGGTGGT

YIM 90564 GGTGGGGTGAAGGCCTACCAAGGCGATTACG ATTCCCCACCTACCGTCAACCCCCAGAGAACTGGGAGCCTGCGTTGGTGGT

YIM 90561 GGTGGGGTGATGGCCTACCAAGGCGACGACG CTTCTCCCGGTACCGTCACTAT------TACGCTTCGTCCCGGGC

YIM 90592 GGTGGGGTGATGGCCTACCAAGGCGACGACG CWTCTCCCGGTACCGTCACTTT------TACGCTTCGTCCCRGGC

YIM 90581 GGTGGGGTGATGGCCTACCAAGGCGACGACG CTTCTCCCGGTACCGTCACTGT------TACGCTTCGTCCCGGGC

YIM 90521 GGTGGGGTGATGGCCTACCAAGGCGACGACG CTTCTCCCGGTACCGTCACTTT------TACGCTTCGTCCCGGGC

YIM 90557 GGTGGGGTGATGGCCTACCAAGGCGACGACG CTTCTCCAGGTACCGTCACCCC------GAAAGGCTTCGTCCCTAGC

YIM 90625 GGTGGGGTGATGGCCTACCAAGGCGGTGACG CTTCTCCAGGTACCGTCAGTAT------CACCCTTCGTCCCTGGC

YIM 90636 GGTGGGGTGATGGCCTACCAAGGCGGTGACG CTTCTCCAGGTACCGTCAGTAT------CACCCTTCGTCCCTGGC Supplementary Table 1. Reference strains used in this study and results obtained from selective PCR amplification with AcpF/AcpR.

Strain PCR result Actinopolyspora halophila DSM 43834T 1 Actinopolyspora mortivallis DSM 44261T 1 Prauserella rugosa DSM 43194T 0 Prauserella alba YIM 90005T 0 Prauserella halophila YIM 90001T 0 Amycolatopsis mediterranei ISP 5501 0 Pseudonocardia zijingensis AS 4.1545T 0 Pseudonocardia ammonioxydans AS 4.1877T 0 Pseudonocardia kongjuensis AS 4.2028T 0 Saccharomonospora halophila DSM 44411T 0 Saccharomonospora paurometabolica YIM 90007T 0 Saccharomonospora xinjiangensis XJ54 0 Saccharopolyspora hirsuta CGMCC 4.1317 0 Thermocrispum municipale IFO 15806 0 Saccharothrix australiensis IFO 14444 0 Lentzea albidocapillata IFO 15855 0 Actinokineospora riparia CCRC 13450 0 Streptomonospora salina YIM 90002T 0 Streptomonospora alba YIM 90003T 0 Nocardiopsis halophila DSM 44494T 0 Nocardiopsis xinjiangensis YIM 90004T 0 Thermobifida alba DSM 43795T 0 Kitasatospora setae CCRC 13404 0 Streptomyces griseus ATCC 23345T 0 Streptomyces thermoviolaceus subsp. thermoviolaceus CCRC 12493 0 Actinocorallia herbida IFO15485 0 Actinomadura madurae CGMCC 4.1178 0 Streptosporangium album CGMCC4.1082 0 Microtetraspora glauca JCM14761 0 Planobispora rosea CCRC 13421 0 Actinoplanes philippinensis JCM 13878 0 Catellatospora citrea subsp. citrea IFO 14495 0 Micromonospora chalcea CGMCC 4.1050 0 Micromonospora yulongensis Y81-917 0 Kribbella alba YIM 31075T 0 Amplification results: 1, band of the expected size; 0, absence of amplification products.