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RAPID IDENTIFICATION OF FILAMENTOUS ACTINOMYCETE GENERA TO THE GENUS LEVEL

RAPID IDENTIFICATION OF FILAMENTOUS ACTINOMYCETE GENERA TO THE GENUS LEVEL. Last updated: 10 th April 2003. Unknown Actinomycete. Sau 3AI. BRANCH B (largest DNA fragment between 980 and 1350 bp ). BRANCH A (largest DNA fragment less than 750 bp ). BRANCH C

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RAPID IDENTIFICATION OF FILAMENTOUS ACTINOMYCETE GENERA TO THE GENUS LEVEL

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  1. RAPID IDENTIFICATION OF FILAMENTOUS ACTINOMYCETE GENERA TO THE GENUS LEVEL Last updated: 10th April 2003

  2. Unknown Actinomycete Sau 3AI BRANCH B (largest DNA fragment between 980 and 1350 bp) BRANCH A (largest DNA fragment less than 750 bp) BRANCH C (2 distinct DNA fragments 300–350 bp & 760–970 bp)

  3. BRANCH A Largest DNA fragment less than 750 bp Sau 3AI digestion Asn I A1 470–590 & 900–960 A2 uncut Kpn I Sca I A1.1 570–630 & 850–900 A1.2 uncut A2.1 410–470 & 1000–1100 A2.2 uncut SphI Sph I Pst I A1.2.1 280–310 & 890–920 A1.2.2 uncut A2.1.1 cut A2.1.2 uncut A2.2.1 400–430 & 1000–1050 A2.2.2 uncut

  4. BRANCH A • Sau 3AI cuts: • largest DNA fragment less than 750 bp • Genera • 1.Glycomyces 3/3 • 2.Gordonia 4/15 • 3.Kitasatospora 12/12 • 4.Microbispora 1/3 • 5.Nocardia 22/27 • 6.Nocardioides 1/5 • 7.Nonomuraea 10/15 • 8.Pseudonocardia 4/17 • 9. Skermania 1/1 • 10.Streptomyces 109/109

  5. A1 Asn I cuts producing 2 bands: 470–590 bp 900–960 bp Genera 1. Glycomyces 3/3 2. Gordonia 4/4 3. Nocardia 22/22 4. Nocardioides 1/1 5. Pseudonocardia 4/4 6. Skermania 1/1 • A2 • Asn I does not cut • Genera • 1.Kitasatospora 12/12 • 2.Microbispora 1/1 • 3.Nonomuraea 10/10 • 4.Sporichthya 2/2 • 5.Streptomyces 109/109

  6. A1.1 • Sca I cuts producing 2 bands: • 570–630 bp • 850–900 bp Species Names: Gordonia (4/15) G. amarae DSM 43392T 1484 G. polyisoprenivorans DSM 44302T 1457 G. rhizosphera IFO 16068T 1475 G. terrae DSM 43249T 1481 Nocardia (22/27) N. abscessus IMMIB D-1592T 1457 N. africana SD769T 1469 N. asteroides ATCC 19247T 1412 N. beijingensis AS4.1521T 1422 N. brasiliensis DSM 43758T 1465 N. brevicatena DSM 43024T 1474 N. carnea DSM 43397T 1472 N. cerradoensis Y9T 1426 Genera 1. Gordonia 4/4 (15) 2. Nocardia 22/22 (27) 3. Skermania 1/1 N. crassostreae NB4HT 1473 N. cyriacigeorgica IMMIB D-1627T 1467 N. farcinica DSM 43665T 1465 N. flavorosea JCM 3332T 1472 N. nova JCM 6044T 1472 N. otitidiscaviarum DSM 43242T 1477 N. paucivorans IMMIB D-1632T 1459 N. pseudobrasiliensis ATCC 51512T 1407 N. pseudovaccinii DSM 43406T 1492 N. seriolae JCM 3360T 1472 N. transvalensis DSM 43405T 1476 N. uniformis JCM 3224T 1472 N. vaccinii DSM 43285T 1472 N. vinacea MK703-102F1T 1476 Skermania (1/1) S. piniformis DSM 43998T 1455

  7. A1.2 • Sca I does not cut • Genera • 1.Glycomyces 3/3 • 2.Nocardioides 1/1 • 3.Pseudonocardia 4/4

  8. A1.2.1 • Sph I cuts producing 2 bands: • 280–310 bp • 890–920 bp • Genera • 1.Glycomyces 3/3 • 2.Nocardioides 1/1 (5) • Species Names: • Glycomyces (3/3) • G. harbinensis IMSNU 20070T 1525 • G. rutgersensis IMSNU 22074T 1522 • G. tenuis IFO 15904T 1489 • Nocardioides (1/5) • N. luteus KCTC 9675T 1469 A1.2.2 Sph I does not cut Genus 1. Pseudonocardia 4/4 (17) Species Names: P. assacharolytica DSM 44247T 1514 P. autotrophica IMSNU 20050T 1512 P. compacta IMSNU 20111T 1510 P. spinosa LM 141T 1512

  9. A2.1 A2.2 • Kpn I cuts producing 2 bands: • 410–470 bp • 1000–1100 bp • Genera • 1.Kitasatospora 12/12 • 2.Sporichthya 2/2 • 3.Streptomyces 109/109 Kpn I does not cut Genera 1.Microbispora 1/1 2.Nonomuraea 10/10

  10. S. echinatus NRRL-ISP 5013T 1448 S. europaeiscabiei CFBP 4497T 1492 S. fimbriatus JCM 4910 1484 S. flavogriseus CBS 101.34T 1518 S. galbus DSM 40089T 1517 S. galilaeus JCM 4757T 1484 S. griseocarneus DSM 40004T 1515 S. griseochromogenes DSM 40499T 1448 S. griseoruber ICSSB 1013T 1436 S. griseus ATCC 10137 1478 S. hygroscopicus ATCC 21431 1517 S. kasugaensis MB 273-C4 1488 S. lanatus NRRL-ISP 5090T 1449 S. lavendulae IFO 12341 1516 S. lipmanii JCM 4711 1484 S. longisporus NRRL-ISP 5166T 1448 S. lydicus ATCC 25470T 1481 S. melanosporofaciens NRRL B-12234T 1483 S. neyagawaensis NRRL-ISP 5588T 1449 S. peucetius JCM 9920T 1486 S. phaeochromogenes DSM 40073T 1450 S. platensis JCM 4662T 1488 S. pseudovenezuelae NRRL-ISP 5212T 1450 S. resistomycificus DSM 40133T 1448 S. reticuliscabiei CFBP 4531T 1448 S. rimosus JCM 4667T 1485 S. scabiei ATCC 25470 1481 S. setonii ATCC 25497T 1532 S. sparsogenes NRRL 2940T 1493 S. stelliscabiei CFBP 4521T 1478 S. subrutilus DSM 40445T 1516 S. tauricus JCM 4837T 1492 S. thermolineatus DSM 41451T 1481 S. turgidiscabies S-27 1528 S. yunnanensis YIM 41004T 1521 S. venezuelae JCM 4526T 1483 S. violaceusniger NRRL-ISP 5563T 1480 S. viridochromogenes JCM 5013 1494 • A2.1.1 • Sph I cuts producing the following bands: • Genera • 1. Kitasatospora 12/12 • 2. Streptomyces 50/109 • Species Names: • Kitasatospora (8/12) [170–190 bp & 1330–1360 bp] • K. azatica IFO 13803T 1481 • K. cheerisanensis YC75T 1508 • K. cineracea SK-3255T 1515 • K. cochleata IFO 14768T 1515 • K. griseola AM-9660T 1517 • K. niigatensis SK-3406T 1516 • K. paracochleata IFO 14769T 1516 • K. phosalacinea KA-338T 1516 • Kitasatospora (4/12) [170–260 bp & 1010–1100 bp] • K. cystarginea IFO 14836T 1523 • K. mediocidica IFO 14789T 1482 • ‘K. melanogena’ JCM 3337 1482 • K. setae KM-6054T 1523 • Streptomyces (50/109) [390–450 bp & 1050–1110 bp] • S. albulus MF 861-C4 1488 • S. argenteolus JCM 4623T 1484 • S. avermitilis NCIMB 12804T 1517 • S. beijiangensis YIM6T 1499 • S. bobili JCM 4624 1483 • S. bottropensis ATCC 25435T 1531 • S. capoamus JCM 4734T 1484 • S. chartreusis NRRL-ISP 5085T 1448 • S. cinnabarinus NRRL-ISP 5467T 1448 • S. clavuligerus JCM 4710T 1486 • S. cyaneus NRRL-ISP 5108T 1448 • S. diastatochromogenes ATCC 12309T 1531

  11. A2.1.2 • Sph I does not cut • Genera • 1.Sporichthya 2/2 • 2.Streptomyces 59/109 • Species Names: • Sporichthya (2/2) • S. brevicatena YU720-21T 1466 • S. polymorpha IFO 12702T 1472 • Streptomyces (59/109) • S. acidiscabies ATCC 49003T 1530 • S. afghaniensis NRRL-ISP 5228 T 1449 • S. albidoflavus DSM 40455T 1476 • S. albofaciens JCM 4342T 1489 • S. albogriseolus NRRL B-1305T 1519 • S. albus subsp. albus JCM 10204 1485 • S. arenae NRRL-ISP 5293T 1447 • S. armeniacus JCM 3070T 1532 • S. bellus NRRL-ISP 5185 T 1450 • S. bikiniensis DSM 40581T 1517 • S. bluensis NRRL-ISP 5564T 1520 • S. caelestis SP 5084T 1448 • S. canescens DSM 40001T 1476 • S. coelicolor DSM 40233T 1476 • S. coeruleofuscus NRRL-ISP 5144 T 1447 • S. coeruleorubidus NRRL-ISP 5145 T 1451 • S. coerulescens NRRL-ISP 5146T 1450 • S. collinus DSM 40129T 1448 • S. eurythermus ATCC 14975T 1531 • S. felleus DSM 40130T 1476 • S. fumanus NRRL-ISP 5154T 1448 • S. glaucescens DSM 40716 1519 • S. gougerotii DSM 40324T 1476 S. griseorubiginosus NRRL-ISP 5469 T 1449 S. hawaiiensis NRRL-ISP 5042 T 1448 S. iakyrus NRRL-ISP 5482T 1448 S. intermedius DSM 40372T 1474 S. janthinus NRRL-ISP 5206T 1448 S. limosus DSM 40131T 1476 S. lincolnensis NRRL 2936T 1519 S. luteogriseus NRRL-ISP 5483T 1448 S. macrosporus DSM 41449T 1484 S. malaysiensis ATB-11T 1475 S. mashuensis DSM 40221T 1518 S. megasporus DSM 41476T 1490 S. mexicanus CH-M-1035T 1450 S. nogalater JCM 4799 1485 S. odorifer DSM 40347T 1476 S. paradoxus MAV 1448 S. pseudogriseolus NRRL B-3985 1516 S. purpurascens JCM 4509T 1484 S. roseoflavus ATCC 19920 1523 S. roseoviolaceus NRRL-ISP 5277 T 1448 S. rutgersensis DSM 40077T 1476 S. sampsonii DSM 40394T 1476 S. scopiformis A25T 1402 S. somaliensis DSM 40738T 1483 S. speibonae DSM 41797T 1490 S. tendae ATCC 19812T 1530 S. thermoalcalitolerans TA56T 1486 S. thermocarboxydovorans DSM 44294 1502 S. thermocarboxydus DSM 44293T 1500 S. thermocoprophilus B19T 1499 S. thermodiastaticus DSM 40573T 1483 S. thermospinosisporus AT10T 1522 S. thermoviolaceus DSM 40443T 1483 S. thermovulgaris DSM 40444T 1486 S. violarus NRRL-ISP 5205 T 1447 S. virginiae ATCC 13013 1415

  12. A2.2.1 • Pst I cuts producing 2 bands: • 400–430 bp • 1000–1050 bp • Genus • 1. Nonomuraea 10/10 (15) • Species Names: • N. africana IFO 14745T 1413 • N. angiospora IFO 13155T 1411 • N. fastidiosa IFO 14680T 1412 • N. helvata IFO 14681T 1411 • N. longicatena NRRL 15532T 1499 • N. polychroma IFO 14345T 1413 • N. recticatena IFO 14525T 1413 • N. roseola IFO 14685T 1414 • N. rubra DSM 43768T 1457 • N. turkmeniaca DSM 43926T 1448 • A2.2.2 • Pst I does not cut • Genus • 1. Microbispora 1/1 (3) • Species Names: • M. rosea IFO 14044T 1473

  13. BRANCH B (Twig 1) largest DNA fragment: 980–1350 bp Sau 3AI digestions B2 410–470 & 900–960 go to BRANCH B (Twig 2) B1 uncut Asn I Kpn I B1.2 410–470 & 1000–1100 B1.1 uncut Sal I/Sph I Sph I B1.1.1 uncut B1.1.2 cut B1.2.2 540 & 910–930 B1.2.1 uncut Pst I Pst I Pst I B1.1.2.1 500–580 & 860–900 B1.1.2.2 uncut B1.2.1.2 cut B1.1.1.1 cut B1.1.1.2 uncut B1.2.1.1 uncut Hind III Sna BI B1.1.1.2.1 520–540 & 890–900 B1.1.1.2.2 uncut B1.2.1.1.1 uncut B1.2.1.1.2 500 & 930

  14. Branch B Sau 3AI cuts: Largest DNA fragment: 980–1300 bp Genera 1. Acrocarpospora 2/2 26. Micromonospora 12/12 2. Actinocorallia 5/5 27. Microtetraspora 3/3 3. Actinokineospora 5/5 28. Nocardia 5/27 4. Actinomadura 28/28 29. Nocardioides 1/1 5. Actinoplanes 21/21 30. Nocardiopsis 13/15 6. Actinopolymorpha 1/1 31. Nonomuraea 5/15 7. Actinosynnema 2/2 32. Oerskovia 3/3 8. Amycolatopsis 12/12 33. Pilimelia 2/2 9. Asanoa 1/1 34. Planomonospora 4/4 10. Catellatospora 3/3 35. Planobispora 2/2 11. Catenuloplanes 2/2 36. Planotetraspora 1/1 12. Couchioplanes 1/1 37. Promicromonospora 2/2 13. Cryptosporangium 4/4 38. Pseudonocardia 12/17 14. Dactylosporangium 6/6 39. Saccharomonospora 2/6 15. Gordonia 11/15 40. Saccharopolyspora 5/8 16. Herbidospora 1/1 41. Saccharothrix 8/8 17. Hongia 1/1 42. Spirilliplanes 1/1 18. Intrasporangium 1/1 43. Spirillospora 2/2 19. Kibdelosporangium 1/1 44. Streptoalloteichus 1/1 20. Kineosporia 4/4 45. Streptomonospora 1/1 21. Kribbella 1/1 46. Streptosporangium 9/12 22. Kutzneria 2/2 47. Thermomonospora 1/2 23. Lechevalieria 2/2 48. Thermobispora 1/1 24. Lentzea 4/4 49. Verrucosispora 1/1 25. Microbispora 2/3 50. Virgisporangium 2/2

  15. B1 • Asn I does not cut • Genera • 1. Acrocarpospora 2/2 11. Nonomuraea 5/5 • 2. Actinocorallia 5/5 12. Planomonospora 4/4 • 3. Actinomadura 28/28 13. Planobispora 2/2 • 4. Actinopolymorpha 1/1 14. Planotetraspora 1/1 • 5. Amycolatopsis 1/12 15. Saccharothrix 1/8 • 6. Cryptosporangium 4/4 16. Spirillospora 2/2 • 7. Herbidospora 1/1 17. Streptomonospora 1/1 • 8. Microbispora 2/3 18. Streptosporangium 9/9 • 9. Microtetraspora 3/3 19. Thermomonospora 1/1 • 10. Nocardiopsis 13/13 20. Thermobispora 1/1

  16. B1.1 • Kpn I does not cut • Genera • 1.Acrocarpospora 2/2 8. Planomonospora 4/4 • 2.Actinomadura 8/28 9. Planobispora 2/2 • 3.Herbidospora 1/1 10. Planotetraspora 1/1 • 4.Microbispora 2/2 11. Streptomonospora 1/1 • 5.Microtetraspora 3/3 12. Streptosporangium 9/9 • 6.Nocardiopsis 13/13 13. Thermobispora 1/1 • 7.Nonomuraea 5/5 14. Thermomonospora 1/1

  17. B1.2 • Kpn I cuts producing 2 bands: • 410–470 bp • 1000–1100 bp • Genera • 1. Actinocorallia 5/5 • 2. Actinomadura 20/28 • 3. Actinopolymorpha 1/1 • 4. Amycolatopsis 1/1 • 5. Cryptosporangium 4/4 • 6. Saccharothrix 1/1 • 7. Spirillospora 2/2

  18. B1.1.1 • Sph I does not cut • Genera • 1. Acrocarpospora 2/2 8. Planomonospora 4/4 • 2. Actinomadura 8/8 9. Planobispora 2/2 • 3. Herbidospora 1/1 10. Planotetraspora 1/1 • 4. Microbispora 2/2 11. Streptosporangium 9/9 • 5. Microtetraspora 3/3 12. Thermomonospora 1/1 • 6. Nocardiopsis 1/13 13. Thermobispora 1/1 • 7. Nonomuraea 5/5 • Sph I cuts producing following bands: • Genera • 1.Nocardiopsis 12/13 • (595–620 bp & 840–860 bp) • 2. Streptomonospora 1/1 • (610–630 bp & 870–900 bp) B1.1.2

  19. Nonomuraea (5/15) N. ferruginea IFO 14094T 1415 N. pusilla IFO 14684T 1414 N. roseoviolacea IFO 14098T 1472 N. salmonea IFO 14687T 1413 N. spiralis IFO 14097T 1409 Planomonospora (4/4) P. alba JCM 9373T 1444 P. parontospora IFO 13880T 1478 P. sphaerica JCM 9374T 1446 P. venezuelensis JCM 3167T 1444 Planobispora (2/2) P. longispora IFO 13918T 1472 P. rosea JCM 3166T 1443 Streptosporangium (8/12) S. amethystogenes DSM 43179T 1449 S. carneum IFO 15562T 1450 S. fragile DSM 43847T 1449 S. longisporum DSM 43180T 1450 S. nondiastaticum DSM 43848T 1444 S. pseudovulgare DSM 43181T 1444 S. viridialbum DSM 43801T 1447 S. vulgare DSM 43802T 1450 Thermomonospora (1/2) T. curvata IFO 15933T 1478 Thermobispora (1/1) T. bispora ATCC 19993T 1469 Nocardiopsis (1/15) N. listeri DSM 40297T 1463 • B1.1.1.1 • Pst I cuts producing 2 bands: • 430–510 bp • 900–1040 bp • Genera • 1.Acrocarpospora 1/2 (2) • 2.Actinomadura 2/8 (28) • 3.Nonomuraea 5/5 (15) • 4.Planomonospora 4/4 • 5.Planobispora 2/2 • 6.Streptosporangium 8/9 (12) • 7.Thermomonospora 1/1 (2) • 8.Thermobispora 1/1 • ( 9. Nocardiopsis 1/1 (15/15) • 590 bp & 870 bp) • Species Names: • Acrocarpospora (1/2) • A. pleiomorpha IFO 16267T 1449 • Actinomadura (2/28) • A. echinospora IFO 14042T 1401 • A. viridis DSM 43175T 1446

  20. B1.1.1.2 Pst I does not cut • Genera • 1.Acrocarpospora 1/2 • 2.Actinomadura 6/8 • 3.Herbidospora 1/1 • 4.Microbispora 2/2 • 5.Microtetraspora 3/3 • 6.Planotetraspora 1/1 • 7.Streptosporangium 1/9

  21. B1.1.1.2.1 • Hind III cuts producing 2 bands: • 520–540 bp • 890–900 bp • Genus • 1. Microtetraspora 3/3 • Species Names: • M. fusca IFO 13915T 1413 • M. glauca IFO 14761T 1475 • M. niveoalba IFO 15239T 1416

  22. B1.1.1.2.2 • Hind III does not cut • Genera • 1.Acrocarpospora 1/1 (2) • 2.Actinomadura 6/6 (28) • 3.Herbidospora 1/1 • 4.Microbispora 2/2 (3) • 5.Planotetraspora 1/1 • 6.Streptosporangium 1/1 (12) • Species Names: • Acrocarpospora (1/2) • A. corrugata DSM 43316T 1449 • Actinomadura (6/28) • A. catellatispora AS 4.1522T 1452 • A. livida JCM 3387T 1466 • A. spadix JCM 3146T 1458 • A. umbrina JCM 6837T 1446 • A. vinacea JCM 3325T 1441 • A. yumaensis JCM 3369T 1454 Herbidospora (1/1) H. cretacea IFO 15474T 1475 Microbispora (2/3) M. corallina DF-32T 1442 M. mesophila JCM 3151T 1456 Planotetraspora (1/1) P. mira IFO 15435T 1475 Streptosporangium (1/12) S. claviforme DSM 44127T 1445

  23. B1.1.2.1 • Pst I cuts producing 2 bands: • 500–580 bp • 860–900 bp • Genus • 1. Nocardiopsis 9/12 (15) • Species Names: • N. alba DSM 43377T 1461 • N. dassonvillei IFO 14626T 1483 • N. exhalans ES10.1T 1454 • N. lucentensis DSM 44048T 1461 • [590 bp & 2 x 400 bp] • N. metallicus R2A 1488 • N. synnemataformans IMMIB D-1215T 1461 • N. tropica VKM Ac-1457T 1452 • N. umidischolae 66/93T 1453 • N. xinjiangensis YIM 90004T 1525 • B1.1.2.2 • Pst I does not cut • Genera • 1.Nocardiopsis 3/12 (15) • 2.Streptomonospora 1/1 • Species Names: • Nocardiopsis (3/15) • N. halotolerans DSM 44410T 1505 • N. kunsanensis HA-9T 1449 • N. trehalosi VKM Ac-942T 1474 • Streptomonospora (1/1) • S. salina YIM 90002T 1529

  24. B1.2.2 • Sal I/Sph I cuts producing 2 bands: • 540 bp • 910–930 bp • Genus • 1.Cryptosporangium 4/4 • Species Names: • C. arvum IFO 15965T 1465 • C. aurantiacum IFO 13967T 1470 • C. japonicum IFO 15966T 1460 • C. minutisporangium IFO 15962T 1470 B1.2.1 Sal I/Sph I does not cut Genera 1.Actinocorallia 5/5 2.Actinomadura 20/20 3.Actinopolyspora 1/1 4.Amycolatopsis 1/1 5.Saccharothrix 1/1 6.Spirillospora 2/2

  25. B1.2.1.1 Pst I does not cut Genera 1.Actinopolymorpha 1/1 2.Amycolatopsis 1/1

  26. B1.2.1.1.1 Sna BI does not cut Genus 1. Actinopolymorpha 1/1 Species Name: A. singaporensis IM 7744T 1442 B1.2.1.1.2 Sna BI cut producing 2 bands: 500 bp & 930 bp Genus 1.Amycolatopsis 1/1 (1/12) Species Name: A. mediterranei DSM 13585 1426

  27. B1.2.1.2 • Pst I cuts: • Genera • 1. Actinocorallia 5/5 • 2. Actinomadura 20/20 (28) • 3. Saccharothrix 1/1 (8) • 4. Spirillospora 2/2 • Species Names: • Actinocorallia (5/5) [400–430 bp & 1000–1050 bp] • A. aurantiaca DSM 43924T • A. glomerata DSM 44360T • A. herbida DSM 44254T • A. libanotica DSM 43554T • A. longicatena DSM 44361T • Actinomadura (4/28) [400–440 bp & 520 bp] • A. madurae DSM 43067T 1445 • A. oligospora ATCC 43269T 1462 • A. rubrobrunea IFO 14622 1438 • A. viridilutea IFO 14480T 1475 Actinomadura (16/28) [400–440 bp & 990–1020 bp] A. atramentaria IFO 14695T 1447 A. citrea IFO 14678T 1447 A. coerulea IFO 14679T 1409 A. cremea IFO 14183 1410 A. fibrosa ATCC 49459T 1450 A. formosensis DSM 43997T 1500 A. fulvescens IFO 14347T 1415 A. hibisca JCM 9627T 1457 A. kijaniata DSM 43764T 1448 A. latina DSM 43382T 1452 A. luteofluorescens IFO 13057T 1466 A. macra IFO 14102T 1403 A. nitritigenes DSM 44137T 1469 A. pelletieri JCM 3388T 1451 A.rugatobispora IFO 14382T 1406 A. verrucosospora IFO 14100T 1411 Saccharothrix (1/8) [465 bp & 990 bp] S. mutabilis DSM 43853T 1458 Spirillospora (2/2) [425–440 bp & 1020–1040 bp] S. albida IFO 12248T 1476 S. rubra JCM 6875T 1445

  28. BRANCH B (Twig 2) B2: 470–590 & 900–960 bp BRANCH B (Twig 1) B2.1 280–310 & 890–920 B2.2 uncut go to Branch B (Twig 3) Sph I Sna BI B2.1.2 470–510 & 970–1000 B2.1.1 uncut Sal I Sal I B2.1.1.1 uncut B2.1.1.2 540–560 & 900–1040 B2.1.2.1 235–265 & 1215–1245 B2.1.2.2 uncut Pvu II Age I Age I B2.1.2.1.1 660–680 & 910–930 B2.1.2.1.2 uncut B2.1.2.2.1 uncut B2.1.1.1.1 560 & 970–980 B2.1.1.1.2 uncut B2.1.2.2.2 cut Sst I/Sac I B2.1.2.1.1.1 585 & 930 B2.1.2.1.1.2 uncut

  29. B2 • Asn I cuts producing 2 bands: • 470–590 bp • 900–960 bp • Genera • 1. Actinokineospora 7/7 18. Lentzea 4/4 • 2. Actinoplanes 21/21 19. Micromonospora 12/12 • 3. Actinosynnema 2/2 20. Nocardia 5/5 • 4. Amycolatopsis 11/12 21. Nocardioides 1/1 • 5. Asanoa 1/1 22. Oerskovia 3/3 • 6. Catellatospora 3/3 23. Pilimelia 2/2 • 7. Catenuloplanes 2/2 24. Promicromonospora 2/2 • 8. Couchioplanes 1/1 25. Pseudonocardia 12/12 • 9. Dactylosporangium 6/6 26. Saccharomonospora 2/2 • 10. Gordonia 11/11 27. Saccharopolyspora 4/4 • 11. Hongia 1/1 28. Saccharothrix 7/8 • 12. Intrasporangium 1/1 29. Spirilliplanes 1/1 • 13. Kibdelosporangium 1/1 30. Streptoalloteichus 1/1 • 14. Kineosporia 4/4 31. Thermocrispum 1/1 • 15. Kribbella 1/1 32. Verrucosispora 1/1 • 16. Kutzneria 2/2 33. Virgisporangium 2/2 • 17. Lechevalieria 2/2

  30. B2.1 • Sph I cuts producing 2 bands: • 280–310 bp • 890–920 bp • Genera • 1. Actinoplanes 21/21 11. Micromonospora 12/12 • 2. Asanoa 1/1 12. Nocardioides 1/1 • 3. Catellatospora 3/3 13. Oerskovia 3/3 • 4. Catenuloplanes 2/2 14. Pilimelia 2/2 • 5. Couchioplanes 1/1 15. Promicromonospora 2/2 • 6. Dactylosporangium 6/6 16. Saccharomonospora 1/4 • 7. Hongia 1/1 17. Spirilliplanes 1/1 • 8. Intrasporangium 1/1 18. Verrucosispora 1/1 • 9. Kineosporia 4/4 19. Virgisporangium 2/2 • 10. Kribbella 1/1 20. Gordonia 3/11 • Species Names: • Gordonia (3/12) [165–175 bp & 1300 bp] • G. alkanivorans DSM 44369T 1490 • G. nitida LE31T 1478 • G. westfalica Kb2T 1478

  31. B2.1.1 • Sna BI does not cut • Genera • 1. Actinoplanes 21/21 7. Micromonospora 12/12 • 2. Asanoa 1/1 8. Pilimelia 2/2 • 3. Catellatospora 3/3 9. Spirilliplanes 1/1 • 4. Catenuloplanes 2/2 10. Verrucosispora 1/1 • 5. Couchioplanes 1/1 11. Virgisporangium 2/2 • 6. Dactylosporangium 6/6

  32. B2.1.1.1 • Sal I does not cut • Genera • 1. Actinoplanes 14/21 • 2. Asanoa 1/1 • 3. Catellatospora 3/3 • 4. Catenuloplanes 2/2 • 5. Dactylosporangium 6/6

  33. Micromonospora (12/12) M. aurantiaca DSM 43813T 1476 M. carbonacea DSM 43168T 1476 M. chalcea DSM 43026T 1477 M. chersina DSM 44151T 1477 M. coerulea DSM 43143T 1476 M. echinospora DSM 43817 1471 M. halophytica DSM 43171T 1476 M. inositola DSM 43819T 1478 M. matsumotoense IMSNU 22003T 1506 M. olivasterospora DSM 43868T 1478 M. purpureochromogenes DSM 43821T 1472 M. rosaria DSM 803T 1477 Pilimelia (2/2) P. anulata DSM 43039T 1468 P. terevasa DSM 43040T 1478 Spirilliplanes (1/1) S. yamanashiensis IFO 15828T 1470 Verrucosispora (1/1) V. gifhornensis HR1-2T 1462 Virgisporangium (2/2) V. aurantiacum YU438-5T 1469 V. ochraceum YU655-43T 1466 • B2.1.1.2 • Sal I cut producing 2 bands: • 540–560 bp • 900–1040 bp • Genera • 1. Actinoplanes 7/21 • 2. Couchioplanes 1/1 • 3. Micromonospora 12/12 • 4. Pilimelia 2/2 • 5. Spirilliplanes 1/1 • 6. Verrucosispora 1/1 • 7. Virgisporangium 2/2 • Species Names: • Actinoplanes(7/21) • A. auranticolor IFO 13992 1475 • A. brasiliensis DSM 43805T 1444 • A. consettensis JCM 7624T 1438 • A. cyaneus IFO 14990T 1469 • A. deccanensis IMSNU 20026T 1452 • A. digitatis IMSNU 22123T 1507 • A. humidus IMSNU 22051T 1470 • Couchioplanes (1/1) • C. caeruleus DSM 44103 1455

  34. B2.1.1.1.1 • Pvu II cuts producing 2 bands: • 560 bp & 970–980 bp • Genus • 1. Dactylosporangium 6/6 • Species Names: • D. aurantiacum DSM 43157T 1476 • D. fulvum DSM 43917T 1475 • D. matsuzakiense DSM 43810T 1469 • D. roseum DSM 43916T 1472 • D. thailandense DSM 43158T 1469 • D. vinaceum DSM 43823T 1476

  35. B2.1.1.1.2 • Pvu II does not cut • Genera • 1. Actinoplanes 14/14 (21) • 2. Asanoa 1/1 • 3. Catellatospora 3/3 • 4. Catenuloplanes 2/2 • Species Names: • Actinoplanes (14/21) • A. campanulatus IFO 12511T 1472 • A. capillaceus K95-5561T 1454 • A. derwentensis IMSNU 20027T 1455 • A. durhamensis IMSNU 22124T 1509 • A. missouriensis DSM 43046T 1484 • A. ferrugineus IMSNU 22125T 1509 • A. globisporus IMSNU 20030T 1452 • A. italicus IFO 13911T 1473 • A. lobatus IFO 12513T 1472 • A. palleronii IFO 14916T 1471 • A. philippinensis IFO 13878T 1472 • A. rectilineatus IFO 13941T 1471 • A. regularis IFO 12514T 1474 • A. utahensis IMSNU 20044T 1508 Asanoa (1/1) A. ferruginea DSM 44099T 1478 Catellatospora (3/3) C. citrea DSM 44098 1478 C. koreensis LM 042T 1510 C. tsunoense DSM 44101T 1452 Catenuloplanes (2/2) C. crispus JCM 9312T 1440 C. japonicus DSM 44102T 1453

  36. B2.1.2 • Sna I cuts producing 2 bands: • 470–510 bp • 970–1000 bp • Genera • 1. Hongia 1/1 • 2. Intrasporangium 1/1 • 3. Kineosporia 4/4 • 4. Kribbella 1/1 • 5. Nocardioides 1/1 • 6. Oerskovia 3/3 • 7. Promicromonospora 2/2 • 8. Saccharopolyspora 1/1

  37. B2.1.2.2 • Sal I does not cut • Genera • 1. Intrasporangium 1/1 • 2. Kineosporia 4/4 • 3. Nocardioides 1/1 • 4. Oerskovia 3/3 • 5. Saccharopolyspora 1/1 B2.1.2.1 Sal I cut producing 2 bands: Genera 1. Hongia 1/1 2. Kribbella 1/1 3. Promicromonospora 2/2

  38. B2.1.2.2.1 Age I does not cut Genera 1. Oerskovia 3/3 2. Saccharopolyspora 1/1 (7) Species Names: Oerskovia (3/3) O. enterophila DSM 43852T 1451 O. jenensis DSM 46097 1496 O. paurometabola DSM 14281T 1486 Saccharopolyspora (1/7) S. thermophila AS4.1511T 1441 • B2.1.2.2.2 • Age I cut producing 2 bands • Genera • 1. Intrasporangium 1/1 • 2. Kineosporia 4/4 • 3. Nocardioides 1/1 (5) • Species Names: • Intrasporangium (1/1) [230 bp & 1240 bp] • I. calvum IFO 12989T 1475 • Kineosporia (4/4) [670–680 bp & 750–830 bp] • K. aurantiaca NRRL B-16913T 1513 • K. rhamnosa JCM 9954T 1474 • K. rhizophila JCM 9960T 1432 • K. succinea JCM 1476 1476 • Nocardioides (1/7) [660 bp & 910 bp] • N. albus DSM 43109T 1450

  39. B2.1.2.1.2 Age I does not cut Genus 1. Promicromonospora 2/2 Species Names: P. citrea DSM 43110T 1453 P. sukumoe DSM 44121T 1460 B2.1.2.1.1 Age I cuts producing 2 bands: 660–680 bp 910–930 bp Genera 1. Hongia 1/1 2. Kribbella 1/1

  40. B2.1.2.1.1.1 • Sst I/Sac I cuts producing 2 bands: • 585 bp • 930 bp • Genus • 1. Hongia 1/1 • Species Name: • H. koreensis IMSNU 50530T 1510 B2.1.2.1.1.2 Sst I/Sac I does not cut Genus 1. Kribbella 1/1 Species Name: K. sandramycini ATCC 39419T 1470

  41. BRANCH 2 (Twig 3) B2.2: uncut BRANCH B (Twig 2) Kpn I B2.2.2 410–470 & 990–1100 B2.2.1 uncut Hind III Hind III B2.2.2.1 uncut B2.2.2.2 cut B2.2.1.1 cut B2.2.1.2 uncut Sal I B2.2.2.1.1 uncut B2.2.2.1.2 540–560 & 950–990 Psp 1406I Psp 1406I B2.2.2.1.1.1 cut B2.2.2.1.1.2 uncut B2.2.2.1.2.1 440 & 1030 B2.2.2.1.2.2 uncut

  42. B2.2 • Sph I does not cut • Genera • 1. Actinokineospora 5/5 9. Nocardia 5/5 • 2. Actinosynnema 2/2 10. Pseudonocardia 12/12 • 3. Amycolatopsis 11/11 11. Saccharomonospora 2/2 • 4. Gordonia 8/11 12. Saccharopolyspora 3/4 • 5. Kibdelosporangium 1/1 13. Saccharothrix 7/7 • 6. Kutzneria 2/2 14. Streptoalloteichus 1/1 • 7. Lechevalieria 2/2 15. Thermocrispum 1/1 • 8. Lentzea 4/4

  43. B2.2.1 Kpn I does not cut Genera 1. Actinosynnema 2/2 2. Gordonia 1/8 3. Kibdelosporangium 1/1 4. Pseudonocardia 3/12 5. Saccharopolyspora 2/3 6. Thermocrispum 1/1 7. Nocardia 5/5 • B2.2.2 • Kpn I cut producing 2 bands: • 410–470 bp • 990–1100 bp • Genera • 1. Actinokineospora 5/5 • 2. Amycolatopsis 11/11 • 3. Gordonia 7/8 • 4. Kutzneria 2/2 • 5. Lechevalieria 2/2 • 6. Lentzea 4/4 • 7. Pseudonocardia 9/12 • 8. Saccharomonospora 2/2 • 9. Saccharopolyspora 1/3 • 10. Saccharothrix 7/7 • 11. Streptoalloteichus 1/1

  44. B2.2.2.1 Hind III does not cut Genera 1. Actinokineospora 5/5 2. Amycolatopsis 11/11 3. Kutzneria 2/2 4. Lechevalieria 2/2 5. Lentzea 1/4 6. Pseudonocardia 8/9 7. Saccharomonospora1/2 8. Saccharopolyspora 1/1 9. Saccharothrix 7/7 10. Streptoalloteichus 1/1 • B2.2.2.2 • Hind III cuts producing 2 bands: • Genera • 1. Gordonia 7/7 (15) • 2. Lentzea 3/4 (4) • 3. Pseudonocardia 1/9 (17) • 4. Saccharomonospora 1/2 (6) • Species Names: • Gordonia (7/15) [170–200 bp & 1270–1290 bp] • G. amicalis IEGMT 1513 • G. bronchialis DSM 43247T 1472 • G. desulfuricans 213ET 1512 • G. hirsuta DSM 44140T 1461 • G. hydrophobica DSM 44015T 1470 • G. namibiensis NAM-BN063AT 1484 • G. rubripertincta DSM 43197T 1485 • Lentzea (3/4) [470–600 bp & 910–970 bp] • L. albidocapillata IMMIB D-958T 1438 • L. violacea IMSNU 50388T 1508 • L. waywayandensis JCM 9114T 1476 • Pseudonocardia (1/17) [210 bp & 1235 bp] • P. sulfidoxydans DSM 44248T 1442 • Saccharomonospora (1/6) [175–185 bp & 980–1005 bp] • S. viridis NCIB 9602T 1490

  45. B2.2.2.1.1 • Sal I does not cut • Genera • 1. Actinokineospora 5/5 • 2. Amycolatopsis 11/11 • 3. Lechevalieria 2/2 • 4. Lentzea 1/1 • 5. Pseudonocardia 8/8 • 6. Saccharomonospora 1/1 • 7. Saccharopolyspora 1/1 • 8. Saccharothrix 7/7 B2.2.2.1.2 Sal I cuts producing 2 bands: 540–560bp 950–990 bp Genera 1. Kutzneria 2/2 2. Streptoalloteichus 1/1

  46. B2.2.2.1.2.1 Psp 1406 I cuts producing 2 bands: 440, 1030 Genus 1. Streptoalloteichus 1/1 Species Name: S. hindustanus IFO 15115T 1474 B2.2.2.1.2.2 Psp 1406 I does not cut Genus 1. Kutzneria 2/2 Species Names: K. kofuensis NRRL B-24061T 1502 K. viridogrisea JCM 3282T 1410

  47. B2.2.2.1.1.1 • Psp 1406I cuts producing 2 bands: • Genera • 1. Amycolatopsis 11/11 (12) • 2. Pseudonocardia 3/8 (17) • 3. Saccharomonospora 1/1 (6) • 4. Saccharopolyspora 1/1 (7) • Species Names: • Amycolatopsis (11/12) [650–680 bp & 740–790 bp] • A. albidoflavus IMSNU 22139T 1474 • A. alba DSM 44262T 1477 • A. azurea IMSNU 20053T 1460 • A. coloradensis DSM 44225T 1483 • A. fastidiosa IMSNU 20054T 1489 • A. methanolica DSM 44096T 1477 • A. orientalis IMSNU 20058T 1458 • A. rubida 13.4T 1402 • A. sacchari DSM 44468T 1484 • A. sulphurea DSM 46092T 1486 • A. thermoflava N1165T 1495 Pseudonocardia (3/17) [700–750 bp & 780 bp] P. alni IMSNU 20049T 1510 P. halophobica IMSNU 21327T 1512 P. hydrocarbonoxydans IMSNU 22140T 1511 Saccharomonospora (1/6) [300 bp; 380 bp & 765 bp] S. xinjiangensis DSM 44391T 1490 Saccharopolyspora (1/7) [380 bp & 765 bp] S. flava AS4.1520T 1443

  48. B2.2.2.1.1.2 • Psp 1406I does not cut • Genera • 1. Actinokineospora 5/5 • 2. Lechevalieria 2/2 • 3. Lentzea 1/1 (4) • 4. Pseudonocardia 5/8 (17) • 5. Saccharothrix 7/7 (8) • Species Names: • Actinokineospora (5/5) • A. diospyrosa NRRL B-24047T 1512 • A. globicatena NRRL B-24048T 1510 • A. inagensis NRRL B-24050T 1507 • A. riparia NRRL B-16432T 1502 • A. terrae IFO 15668T 1471 • Lechevalieria (2/2) • L. aerocolonigenes NRRL B-16139 1499 • L. flava NRRL B-16131T 1510 • Lentzea (1/4) • L. albida 37-28 1436 Pseudonocardia (5/17) P. alaniniphila CCTCC AA97001T 1557 P. aurantiaca CCTCC AA97002T 1556 P. thermophila IMSNU 20112T 1520 P. yunnanensis CCTCC M 90959T 1555 P. zijingensis AS 4.1545T 1422 Saccharothrix (7/8) S. australiensis NRRL 11239T 1413 S. coeruleofusca NRRL B-16115T 1512 S. espanaensis NRRL 15764T 1510 S. longispora NRRL B-11616T 1456 S. syringae NRRL B-16468T 1510 S. tangerinus MK27-91F2T 1475 S. texasensis NRRL B-16134T 1512

  49. B2.2.1.1 • Hind III cuts producing 2 bands: • Genera • 1. Gordonia 1/1 (15) • 2. Kibdelosporangium 1/1 • 3. Nocardia 5/5 (27) • Species Names: • Gordonia (1/15) [170–200 bp & 1270–1290 bp] • G. sputi DSM 43896T 1486 • Kibdelosporangium (1/1) [460 bp & 990 bp] • K. aridum 1450 • Nocardia (5/27) [550–570 bp & 890–990 bp] • N. cummidelens R89T 1458 • N. fluminea S1T 1462 • N. ignorata DSM 44496T 1489 • N. salmonicida JCM 4826T 1474 • N. soli W30T 1466

  50. B2.2.1.2 • Hind III does not cut • Genera • 1. Actinosynnema 2/2 • 2. Pseudonocardia 3/3 (17) • 3. Saccharopolyspora 2/2 (7) • 4. Thermocrispum 1/1 (2) • Species Names: • Actinosynnema (2/2) • A. mirum DSM 43827T 1460 • A. pretiosum subsp. pretiosum NRRL B-16060T 1504 • Pseudonocardia (3/17) • P. petroleophila IMSNU 22072T 1515 • P. saturnea IMSNU 20052T 1519 • P. xinjiangensis CCTCC AA97020T 1563 • Saccharopolyspora (2/7) • S. hirsuta ATCC 27875T 1487 • S. spinosa DSM 44228T 1465 • Thermocrispum (1/2) • T. agreste DSM 44070T 1518

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