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126

Chapter 3

Appendix

Chapter 3

Fe.o Hy.r;Ta.v Ag.c;Se.j Br.m

Pl.l Ac.m Ho.l; Le.v An.o Lo.c

Figure A1. PCR-DGGE with universal bacterial primers for the rhizosphere of 12 different plant species. Fe.oFestuca ovin; Hy.r- Hypochaeris radicata; Ta.v- Tanacetum vulgareI; Ag.c- Agrostis capillaries; Se.j- Senecio jacobaea; Br.m- Briza media; Pl.l- Plantago lanceolata; Ac.m- Achillea millefolium; Ho.l- Holcus lanatus; Le.vLeucanthemum vulgare; An.o- Anthoxanthum odoratum; Lo.c- Lotus corniculatus.

127

Appendix

Figure A2. Acidobacteria - phylum specific PCR-DGGE.

Fe.o Hy.r;Ta.v Ag.c;Se.j Br.m

Pl.l Ac.m Ho.l Le.v An.o Lo.c

Figure A3. Verrucomicrobia - phylum specific PCR-DGGE for the rhizosphere of 12 different plant species. Fe.oFestuca ovin; Hy.r- Hypochaeris radicata; Ta.v- Tanacetum vulgareI; Ag.c- Agrostis capillaries; Se.j- Senecio jacobaea; Br.m- Briza media; Pl.l- Plantago lanceolata; Ac.m- Achillea millefolium; Ho.l- Holcus lanatus; Le.vLeucanthemum vulgare; An.o- Anthoxanthum odoratum; Lo.c- Lotus corniculatus.

128

Table A 1. Identified ribosomal genes and predicted ORF of the acidobacterial genomic fragment 59

1

Locus

Gene Length

Best Match Accession nr.

1-651

650

AAP58487

2

651 -1685

1034

AAP58486

3

1682 -2296

614

AAP58484

4

2296 -3084

788

YP001432775

5

3926 -3081

845

AAP58482

3945 -5228

1283

AAP58586

6 7 8 9 10

12319 11606 12328 12405 12443 13459 13991 13830

Source organism uncultured Acidobacteria bacterium; Acidobacteria uncultured Acidobacteria bacterium; Acido bacteria uncultured Acidobacteria bacterium; Acidobacteria Roseiflexus castenholzii DSM 13941; Chloroflexi uncultured Acidobacteria bacterium; Acidobacteria uncultured Acidobacteria bacterium; Acidobacteria

Description

Identity (%)

E value

hypothetical protein

48%

5.00E -48

putative phosphoribosylformylglycinamidine cyclo ligase

67%

8.00E -133

putative trifunctional purine biosynthesis protein

68%

3.00E -70

methyltransferase type 12

38%

4.00E -37

conserved hypothetical protein

56%

1.00E -81

putative tyrosyl -tRNA synthetase

60%

9.00E -145

protein of unknown function DUF583

40%

4.00E -15

beta-lactamase

48%

2.00E -94

peptide methionine s ulfoxide reductase msrB

72%

1.00E -49

hypothetical protein Acid_7376

36%

1.00E -134

713 77 1016 161

11

14424 14774

350

YP745980

12

16187 15102

1085

YP828570

13

16292 16717

425

YP591253

14

16850 20080

3230

YP593086

Thioalkalivibrio sp. HL EbGR7; Gammaproteobacteria Acidobacteria bacterium Ellin345; Acidobacteria Granulibacter bethesdensis CGDNIH1; Alphaproteobacteria Solibacter usitatus Ellin6076; Acidobacteria

Chapter 5

Chapter 5

129

16 17 18 19 20 21 22 23 24 25 26 27

20185 21 181 21 230216 07 21 912 21 736 24 963 22 045 271 8125 124 27 213 28 001 28 2892887 34 29000 31 393 31 715 32362 32 401 32 715 33 079 33 558 33723 36200 36197 >37657

Appendix

130

15

9 96 378 1 77 2 919

YP823226

2 058

YP476496

7 89

YP002753331

5 85

ZP01884371

2 394

YP002753332

Solibacter usit atus Ellin6076; Acidobacteria Solibacter usitatus Ellin6076; Acidobacteria Solibacter usit atus Ellin6076; Acidobacteria Synechococcus sp. JA -23B'a(2 -13); Bacteroidetes Acidobacterium capsulatum ATCC 51196; Acidobacteria

BNR repeat-containing glycosyl hydrolase

38%

3.00E -158

peptidase S9 prolyl oligopeptidase

31%

1.00E -76

glyoxalase/bleomycin resistance protein/dioxygenase

42%

5.00E -51

methionine -S-sulfoxide reductase

71%

8.00E -69

ATP -dependent helicase HrpB

36%

2.00E -109

permease

33%

3.00E -126

FG-GAP repeat-containing protein

44%

8.00E -93

6 48 3 15 4 80 2 478

YP001816913

14 61

YP758955

Opitutus terrae PB90 Verrucomicrobium Hyphomonas neptunium ATCC 15444; Alphaproteobacteria

-1;

101 -28

73

tRNA Type Ile Anti Codon GAT

592 6-7403 8144 11130 11517 11401

1477

16S rRNA

2986

23S rRNA

116

5S rRNA

Table A 2. Identified ribosomal genes and predicted ORF of the acidobacterial genomic fragment 66 Gene Match Accession nr.

Locus

Length

1

298 -2

297

2

789 -295

495

ACB85718

3

2986 -782

2205

YP001616538

4 5 6 7 8 9 10 11 12 13 14 15 16

4000 3080 4828 4019 6439 5042 7455 6538 7597 8160 8602 8204 8701 10572 10579 11775 12197 11844 13143 12511 13276 14151 14334 14546 15416 14688

Source organism

Natranaerobius thermop hilus JW/NM -WN-LF; Firmicutes Sorangium cellulosum 'So ce 56'; Firmicutes

Description

Identity (% )

E value

(2Fe-2S) -binding domain protein

42%

3.00E -28

putative xanthine dehydrogenase

48%

0

putative neutral zinc metallopeptidase.

50%

4.00E -60

M ATE efflux family protein

22%

2.00E -37

Glyoxalase/bleomycin resistance protein/dioxygenase

45%

4.00E -28

response regulator

38%

1.00E -15

oligoendopeptidase F

48%

2.00E -164

beta-lactamase

52%

5.00E -100

921 810

YP001618097

1398

YP001918496

918

ZP03131181

Sorangium cellulo sum 'So ce 56'; Firmicutes Natranaerobius thermophilus JW/NM -WN-LF; Firmicutes Chthoniobacter flavus Ellin428; Verrucomicrobia

564 399

YP628793

1872

YP001378361

1197

YP593265

M yxococcus xanthus DK 1622; Deltaproteobacteria Anaeromyxobacter sp. Fw109 -5; Deltaproteobacteria Acidobacteria bacterium Ellin345; Acidobacteria

354 633

AAK44006

Arabidopsis t haliana

unknown protein

42%

3.00E -37

876

YP822718

Solibacter usitatus Ellin6076; Acidobacteria

hypothetical protein Acid_1440

42%

6.00E -50

YP826143

Solibacter usitatus Ellin6076; Acidobacteria

hypothetical protein Acid_4899

46%

1.00E -48

213 729

Chapter 5

131

18 19 20

15742 16302 16409 16522 16770 16600 29899 23915 2115 221080 21347 21274 21497 22944 17519 20515 16987 17103

Appendix

132

17

561 114 171 5985

YP001231010

Geobacter uraniireducens Rf4; Deltaproteobacteria

PKD domain-containing protein

72

tRNA Type Ala Anti Codon TGC

73

tRNA Type Ile Anti Codon GAT

1447

16S rRNA

2996

23S rRNA

116

5S rRNA

41%

2.00E -153

Table A 3. Identified ribosomal genes and predicted ORF of the aci dobacterial genomic fragment 70

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16

Locus

Gene Length

1122 -196

926

2649 1279 3559 2675 4254 3556 4463 5437 5509 6828 7515 6859 8547 7600 9380 8544 9435 10679 17552 17974 18287 17985 19046 18306 19281 19628 19631 22282 2363622314

Best Match Accession nr.

1370

ZP01907632

884

ZP01907633

698

YP001611503

974

YP001232870

Source organism

Plesiocystis pacifica SIR -1; Deltaproteobacteria Plesiocystis pacifica SIR -1; Deltaproteobacteria Sorangium cellulosum 'So ce 56'; Deltaproteobacteria Geobacter uraniumreducens Rf4; Deltaproteobacteria

Description

Identity (% )

E value

type I phosphodiesterase/nuc leotide pyrophosphatase

58%

1.00E -154

hydrolase, TatD family protein

62%

2.00E -106

hypothet ical protein sce0866

47%

2.00E -39

O-methyltransferase, family 2

34%

3.00E -47

conserved hypothetical protein

64%

2.00E -94

putative tyrosyl -tRNA synthetase

61%

1.00E -141

hypothetical protein Acid_3039

74%

5.00E -51

hypothetical protein PphaDRAFT_2152

27%

4.00E -14

transcriptional regulator, PadR -like family

51%

3.00E -28

protein of unknown function DUF214

34%

6.00E -127

protein of unknown function DUF1552

41%

1.00E -96

1319 656 947 836

AAP58482

1244

AAP58586

422

YP824303

uncultured Acidobacteria bacterium; Acidobacteria uncultured Acidobacteria bacterium; Acidobacteria Solibacter usitatus Ellin6076; Acidobacteria

302 740

ZP00589530

347

YP823582

2651

YP826039

1322

YP823307

Pelodictyon phaeoclathratiforme BU -1; Chlorobi Solibacter usitatus Ellin6076; Acidobacteria Solibacter usitatus Ellin6076; Acidobacteria Solibacter usitatus Ellin6076; Acidobacteria

Chapter 5

133

18 19 20 21 22 23 24 25

26

26018 23652 27570 26020 27660 28160 28170 28664 28658 29986 29979 30611 30608 32272 32269 32757 32829 33227

2366

YP827058

1550

XP001181739

500

ZP01061645

494

NP662719

1328

YP824740

632

YP590766

1664

YP590765

488

YP593001

398

ZP02057379

33224 33757 13424 13500 13710 13782 11754 13299 14097 17089 17311 17427

533

YP321699

Solibacter usitatus Ellin6076; Acidobacteria Strongylocentrotus purpuratus; M etazoa Flavobacterium sp. M ED217; Bacteroidetes Chlorobium tepidum TLS; Chlorobi Solibacter usitatus Ellin6076; Acidobacteria Acidobacteria bacterium Ellin345; Acidobacteria Acidobacteria bacterium Ellin345; Acidobacteria Acidobacteria bacterium Ellin345; Acidobacteria M ethylobacterium chloromethanicum CM 4; Alphaproteobacteria Anabaena variabilis ATCC 29413; Cyanobacteria

Protein of unknown function DUF1592

45%

0

PREDICTED: similar to ankyrin 2,3/unc44

30%

1.00E -44

hypothetical protein M ED217_08675

49%

8.00E -36

hypothetical protein CT1840

36%

3.00E -18

integral membrane sensor signal transduction histidine kinase two component transcriptional regulator, LuxR family

56%

3.00E -127

67%

7.00E -69

Integrin-like protein

66%

0

Thioredoxin -like

39%

2.00E -25

methionine -R -sulfoxide reductase

72%

2.00E -47

methionine sulfoxide reductase A

76%

4.00E -77

76

tRNA Type Ile Anti Codon GAT

72

tRNA Type Ala Anti Codon TGC

1545

16S rRNA

2992

23S rRNA

116

5S rRNA

Appendix

134

17

Table A 4. Identified ribosomal genes and predicted ORF of the acidobacterial genomic fragment 89 Locus 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16

3373 3963 5684 4431 7550 5685 8606 7626 9652 8603 10001 9639 10977 9982 11016 13352 14760 13906 14895 15623 15654 16481 17780 16491 19021 17741 20023 19031 20049 21266 22161 21703

Gene Length

Best Match Accession nr.

591

YP822288

1254

YP826794

1866

YP826795

981

YP826177

1050

YP826178

363

YP826179

996 2337

YP826179 YP826180

Source organism Solibacter usitatus Acidobacteria Solibacter usitatus Acidobacteria Solibacter usitatus Acidobacteria Solibacter usitatus Acidobacteria Solibacter usitatus Acidobacteria Solibacter usitatus Acidobacteria Solibacter usitatus Acidobacteria Solibacter usitatus Acidobacteria

Ellin6076; Ellin6076; Elli n6076; Ellin6076; Ellin6076; Ellin6076; Ellin6076; Ellin6076;

Description

Identity (% )

E value

hypothetical protein Acid_1005

0.62

4E-39

hypothetical protein Acid_5562

0.58

4E-130

hypothetical protein Acid_5563

0.53

3E-178

hypothetical protein Acid_4933

0.55

6E-87

hypot hetical protein Acid_4934

0.45

6E-79

hypothetical protein Acid_4935

0.67

1E-35

hypothetical protein Acid_4935

0.62

7E-95

penicillin amidase

0.68

0

polysaccharide deacetylase

0.31

1E-19

M scS mechanosensitive ion channel

0.65

2E-98

glycosyl transferase, group 1

0.31

7E-24

major facilitator transporter

0.58

9E-135

tryptophanyl -tRNA synthetase

0.77

4E-142

23S rRNA (uracil -5 -)-methyltransferase RumA

0.54

4E-119

GCN5-related N-acetyltransferase

0.6

2E-43

855 729

YP644691

828

YP823555

1290

ZP01516089

1281

YP827069

993

YP827070

1218

YP827071

459

YP561875

Rubrobacter xylanophilus DSM 9941; Actinobacteria Solibacter usitatus Ellin6076; Acidobacteria Chloroflexus aggregans DSM 9485; Chloroflexi Solibacter usitatus Ellin6076; Acidobacteria Solibacter usitatus Ellin6076; Acidobacteria Solibacter usitatus Ellin6076; Acidobacteria Shewanella denitrificans OS217; Gammaproteobacteria

Chapter 5

135

18 19 20 21 22 23 24 25

24233 22164 25493 24252 26528 25497 27023 26544 28664 27177 29006 29428 29960 29475 31307 29997 31618 31436 1225 1150 1116 1040 1422 2917 2-876 931-1049

2070

YP823101

1242

YP822843

1032

YP822844

480

YP822845

1488

YP822847

Solibacter usitatus Ellin6076; Acidobacteria Solibacter usitatus Ellin6076; Acidobacteria Solibacter usitat us Ellin6076; Acidobacteria Solibacter usitatus Ellin6076; Acidobacteria Solibacter usitatus Ellin6076; Acidob acteria

peptidase S9 prolyl oligopeptidase

0.55

0

competence/damage -inducible protein CinA

0.69

6E-155

hypothetical protein Acid_1568

0.75

1E-152

phosphatidylglycerophosphatase

0.64

2E-47

M iaB-like tRNA modifying enzyme YliG

0.81

0

hypothetical protein Acid_5811

0.55

4E-30

AAA ATPase

0.57

7E-128

423 486

YP827043

1311

YP829028

Solibacter usitatus Ellin6076; Acidobacteria Solibacter usitatus Ellin6076; Acidobacteria

183 75

tRNA Type Ala Anti Codon TGC

76

tRNA Type Ile Anti Codon GAT 16S rRNA 23S rRNA 5S rRNA

Appendix

136

17

Table A 5. Identified ribosomal genes and predicted ORF of the acidobacterial genomic fragment 92 Gene Best Match Accession nr.

Locus

Gene Length

1

1 -270

270

2

448 -1323

876

ZP01665324

3

1333 -1875

543

YP843383

4

2684 -1941

744

CAO90041

5

3276 -2677 10289 12088 13322 12108 13376 14203 14564 14205 14911 14561 16383 15226 19085 16818 19801 19106 20811 19798 21007 21363 21507 21683 21822 22040

6 7 8 9 10 11 12 13 14 15 16 17

600

YP674542

1800

YP721564

1215

AAP58481

828

AAP58482

Source organism

Thermosinus carboxydivorans Nor1; Firmicutes Methanosaeta thermophila PT; Archaea; Euryarchaeota Microcystis aeruginosa PCC 7806 ; Cyanobacteria Trichodesmium erythraeum IMS101; Cyanobacteria uncultured Acidobacteria bacterium; Acidobacteria uncultured Acidobacteria bacterium; Acidobacteria

Description

Identity (%)

E value

glucose-1-phosphate thymidylyltransferase

64%

7.00E-102

dTDP-4-dehydrorhamnose 3,5 -epimerase

61%

1.00E-59

unnamed protein product

38%

1.00E-32

peptidyl -Asp metallopeptidase. metallo peptidase MEROPS family M72

34%

9.00E-24

putative tyrosyl-tRNA synthetase

66%

2.00E-143

conserved hypothetical protein

55%

7.00E-77

TPR repeat-containing protein

27%

2.00E-67

eight transmembrane protein EpsH

40%

1.00E-30

eight transmembrane protein EpsH

37%

1.00E-36

360 351 1158 2268

YP824583

696

YP780028

1014

ZP03022916

Solibacter usitatus Ellin6076; Acidobacteria Rhodopseudomonas palustris BisA53; Alphaproteobacteria Geobacter sp. M21; Deltaproteobacteria

357 177 219 Chapter 5

137

19 20 21 22

22083 22481 22515 22940 27686 23364 28184 28444 28612 33279

399

XP001917939

426

YP825373

4323

YP827784

261

YP001799707

4668

YP873691

Appendix

138

18

Solibacter usitatus Ellin6076; Acidobacteria

hypothetical protein Acid_6577

33%

2.00E-56

Acidothermus cellulolyticus 11B; Actinobacteria

hypothetical protein Acel_1933

52%

0

7726 -7650

76

7445 -7373

72

tRNA Type Ile Anti Codon GAT tRNA Type Ala Anti Codon TGC

7891 -9436

1545

16S rRNA

4063 -7072

3009

23S rRNA

3668 -3784

116

5S rRNA

Table A6. Identified ribosomal genes and predicted ORF of the acidobacterial genomic fragment 98 Locus

Gene Length

Best Match Accession nr.

1

488-45

444

YP113754

2

1185 -463

723

YP002762714

402

YP002762713

1290

YP002762713

3 1 2 3

1571 1170 2706 1416 3064 2747 5017 3713

Source organism M ethylococcus capsulatus str. Bath; Gamm aproteobacteria Gemmatimonas aurantiaca T -27; Gemmatimonadetes Gemmatimonas aurantiaca T -27 ; Gemmatimonadetes Gemmatimonas aurantiaca T -27; Gemmatimonadetes

Description

Identity (% )

E value

hypothetical protein M CA1292

29%

0.004

51%

2.00E -55

78%

9.00E -53

41%

1.00E -67

amine oxidase, flavin -containing

43%

2.00E -67

UbiE/COQ5 methyltransferase

48%

1.00E -61

putative fapy -DNA glycosylase

66%

2.00E -114

hypothetical protein BBR47_24320

30%

2.00E -22

putative carboxy -terminal processing protease

53%

3.00E -123

putative tyrosyl -tRNA synthetase

63%

1.00E -150

conserved hypothetical protein

66%

7.00E -91

putative trifunctio nal purine biosynthesis protein

69%

3.00E -67

putative amino acid ABC transporter ATP -binding protein putative amino acid ABC transporter permease/substrate binding protein putative amino acid ABC transporter permease/substrate binding protein

317 1304

YP628484

5069 5848

779

YP590824

5

6803 5904

899

AAP58579

6

7544 6807

737

YP002771913

7

8097 7507

590

8

9603 8296

1307

AAP58520

9

17261 15924

1337

AAP58586

10

17286 18110

824

AAP58482

11

18694 18107

587

AAP58484

uncultured Acidobacteria bacterium; Acidobacteria uncultured Acidobacteria bacterium; Acidobacteria uncultured Acidobacteria bacterium; Acidobacteria uncultured Acidobacteria bacterium;

139

Chapter 5

4

M yxococcus xanthus DK 1622; Deltaproteobacteri Acidobacteria bacterium Ellin345; Acidobacteria uncultured Acidobacteria bacterium; Acidobacteria Brevibacillus brevis NBRC 100599; Firmicutes

19802 18771

1031

AAP58588

13

20982 19852

1130

AAP58487

14

21460 21020

440

AAP58589

15

22502 21447

1055

AAP58589

16

24839 22599

2240

AAP58590

17

25585 24878

707

AAP58591

18

25848 25585

263

AAP58592

19

27226 25871

1355

AAP58593

20

28560 27301

1259

AAP58594

21

28774 29850

1076

AAP58595

22

30793 29807

986

AAP58596

13619 13543 13404 13329

uncultured Acidobacteria bacterium; Acidobacteria uncultured Acidobacteria bacterium; Acidobacteria uncultured Acidobacteria bacterium; Acidobacteria uncultured Acidobacteria bacterium; Acidobacteria uncultured Acidobacteria bacterium; Acidobacteria uncultured Acidobacteria bacterium; Acidobac teria uncultured Acidobacteria bacterium; Acidobacteria uncu ltured Acidobacteria bacterium; Acidobacteria uncultured Acidobacteria bacterium; Acidobacteria uncultured Acidoba cteria bacterium; Acidobacteria uncultured Acidobacteria bacterium; Acidobacteria

putative cycloligase

71%

5.00E -139

hypothetic al protein

39%

6.00E -61

putative amidophosphoribosyl transferase

76%

1.00E -54

putative amidoph osphoribosyl transferase

89%

4.00E -153

putative phosphoribosylformylglycinamidine synthase II

76%

0

putative phosphoribosylformylglycinamidine synthase I

75%

7.00E -101

putative phosphoribosylformylglycinamidine synthase I

78%

4.00E -27

putative adenylosuccinate lyase

71%

6.00E -172

putative protease

75%

4.00E -180

conserved hypothetical protein

79%

9.00E -152

conserved hypothetical protein

60%

6.00E -95

76

tRNA Type Ile Anti Codon GAT

75

tRNA Type Ala Anti Codon TGC

Appendix

140

12

13849 15392 10041 13030 9747 9863

1543

16S rRNA

2989

23S rRNA

116

5S rRNA

Chapter 5

141

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16

Locus

Gene Le ngth

Best Match Accession nr.

1539 -