TABLE 2 Oligonucleotide primers and probes used in FISH* S1PRO634RHLIProchlorococcus GCC GAT CAG TTT CCA CTG S2PRO634RHLIIProchlorococcus GCC TTT CAG.

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TABLE 2 Oligonucleotide primers and probes used in FISH* S1PRO634RHLIProchlorococcus GCC GAT CAG TTT CCA CTG S2PRO634RHLIIProchlorococcus GCC TTT CAG TTT CCA CTG DPRO634RLL Prochlorococcus (PAC1/NATL1-MIT cluster) GCC AAT CAG TTT CCA CTG. Oligonucleotide ProbesSpecific target DNASequence 5´-3´ 645 HLI*HLIProchlorococcus ACC ATA CTC AAG CCG ATC 645 HLII*HLII Prochlorococcus ACC ATA CTC AAG CCT TTC 645 LL*LL Prochlorococcus (PAC1/NATL1-MIT cluster) ACC ATA CTC AAG CCA ATC 181 HLI*HLI Prochlorococcus CAT TTC ACC TAT CGG CAT 181 LL*LL Prochlorococcus (not MIT9211 & includes some Syns) CAT TTC ACC TCT CGG CAT 148 LL-MIT*LL Prochlorococcus (MIT9211,MIT9303, MIT9313 only) CCG TTT CCA ACC GTT ATC 405PRO*Gen Prochlorococcus (General, but not MIT9303, MIT9313) AGA GGC CTT CGT CCC TCA

TABLE 1 Oligonucleotide primers and probes used in dot blots and DGGE analysis PCR PrimersSpecific target DNASequence 5´-3´ OXY107F*Oxygenic phototroph biased GGA CGG GTG AGT AAC GC GTG R OXY1313R*Oxygenic phototroph biased CTT CAC GTA GGC GAG TTG CAG C CYA359F†Oxygenic phototroph specific GGG GAA TYT TCC GCA ATG GG PRO1017RProchlorococcus, excluding MIT9303, MIT9313, MIT9302 TCC CGA AGG CAC CCT CWA AA 27FEubacterial AGA GTT TGA TCM TGG CTC AG 1224REubacterial CAT TGT AGC ACG TGT GTA Oligonucleotide ProbesSpecific target DNASequence 5´-3´ S1PRO634RHLIProchlorococcus GCC GAT CAG TTT CCA CTG S2PRO634RHLIIProchlorococcus GCC TTT CAG TTT CCA CTG DPRO634RLLProchlorococcus (except SS120) GCC AAT CAG TTT CCA CTG SARG634RSS120 GCC CTT CAG TTT CCA CTG MIT1023RMIT9303 TGC GTT CCC AAA GGC ACT EUB338Eubacteria GCT GCC TCC CGT AGG AGT PRO444R ‡Prochlorococcus, excluding MIT9303, MIT9313 TAT TCC TCA AGT ACC GTC ATA MAR572RMarine picophytoplankton clade biased GCC GCC TGC GGA CGC TTT † A GC-clamp consisting of a 40 nucleotide GC-rich sequence (5´-CGC CCG CCG CGC CCC GCG CCG GTC CCG CCG CCC CCG CCC-3´) was attached to the 5´ end of the primer and used in DGGE analysis. ‡ Based on SAR6R

TABLE 2 Oligonucleotide primers and probes used in FISH* S1PRO634RHLIProchlorococcus GCC GAT CAG TTT CCA CTG S2PRO634RHLIIProchlorococcus GCC TTT CAG TTT CCA CTG DPRO634RLL Prochlorococcus (PAC1/NATL1-MIT cluster) GCC AAT CAG TTT CCA CTG. Oligonucleotide ProbesSpecific target DNASequence 5´-3´ 645 HLI*HLIProchlorococcus ACC ATA CTC AAG CCG ATC 645 HLII*HLII Prochlorococcus ACC ATA CTC AAG CCT TTC 645 LL*LL Prochlorococcus (PAC1/NATL1-MIT cluster) ACC ATA CTC AAG CCA ATC 181 HLI*HLI Prochlorococcus CAT TTC ACC TAT CGG CAT 181 LL*LL Prochlorococcus (not MIT9211 & includes some Syns) CAT TTC ACC TCT CGG CAT 148 LL-MIT*LL Prochlorococcus (MIT9211,MIT9303, MIT9313 only) CCG TTT CCA ACC GTT ATC 405PRO*Gen Prochlorococcus (General, but not MIT9303, MIT9313) AGA GGC CTT CGT CCC TCA

181LL 645HLI 181HLI 645LL 1445NATL 645HLII 405PRO 405SYN 0.1 Prochlorococcus sp. MIT9107 Prochlorococcus sp. SB Prochlorococcus sp. TAK Prochlorococcus sp. MIT9215 Prochlorococcus sp. MIT9202 Prochlorococcus sp. MIT9201 Prochlorococcus sp. MIT9302 Prochlorococcus sp. GP2 Prochlorococcus sp. MIT9312 Prochlorococcus sp. EQPAC1 Prochlorococcus sp. PCC9511 Prochlorococcus sp. CCMP1378 Prochlorococcus sp. NATL2A Prochlorococcus sp. NATL2B Prochlorococcus sp. PAC1 Prochlorococcus sp. NATL1MIT Prochlorococcus marinus SS120 Prochlorococcus sp. MIT9211 Prochlorococcus sp. MIT9303 Prochlorococcus sp. MIT9313 Synechococcus sp. WH8018 Synechococcus sp. WH7805 Synechococcus sp. WH7803 Synechococcus sp. WH8103 Synechococcus sp. WH8101 Synechococcus sp. PCC6307 Synechococcus sp. PCC LL

Eastern North Atlantic –stratified HLI and LL genotypes correlated with mixed layer depth  in agreement with clone libraries, DGGE analysis and FISH data Western North Atlantic –uniform distribution of HLII and LL genotypes in a mixed water column Mediterranean Sea –stratified HLI and LL genotypes as observed in the eastern North Atlantic Red Sea –stratified HLII and LL genotypes Distribution of Prochlorococcus genotypes HLII LL HLI LL HLI LL HLII LL

Location/Depth (m) Relative hybridisation (%) HLILLHLIISARG AM2 15 m (20.4°S, 70.6°W)* AM1 55 m (20.4°S, 70.6°W)** * above the oxycline ** below the oxycline 15m m m m m m m (?) m Prochlorococcus dot blot data : Coastal Chile (20.4°S, 70.6°W) low oxygen environment

oligotophy/eutrophy gradient of surface waters along the transect oxygen-depleted (ca 2-10 µM O 2 ) waters that occur at depths from ca metres throughout the basin. divergent driven, gentle but continual upwelling of the Equator and adjacent downwelling waters Indian Ocean - Arabian Sea - cruise Sept 2001

Marine Synechococcus strains: Roscoff StrainOriginIsolatorLocationPUB:PEB Minos 1 Minos 2 Minos 11 Minos 12 Max 42 Almo 3 Eum 14 Oli 31 Mediterranean Sea Sargasso Sea Alboran Sea Tropical Atlantic Equatorial Pacific F. Partensky D. Vaulot 39°10’N 6 °10’E 34°0’N 18 °10’E 26°18’N 63°26’W 36°11’N 1°51’W 21°02’N 31°0’W 5°N 150° 0°W PEB>PUB PEB<PUB

Analysis of Synechococcus strains III : Red Sea StrainDate of Depth of Colour PUB/PEB MotilityN sourcentcA RFLP16S sequence isolationisolation ratio type RS mRed 0.7 -NO 3 - /NH 4 + I + RS mOrange 1.4 -NO 3 - /NH 4 + II + RS mOrange 2.0 -NO 3 - /NH 4 + III + RS mOrange 1.5 -NO 3 - /NH 4 + III + RS mOrange/Red (+)NO 3 - /NH 4 + IV + RS mGreen - -NO 3 - /NH 4 + N/A + RS mRed 0.5 -NO 3 - /NH 4 + II + RS mRed 0.5 -NO 3 - /NH 4 + II + RS mGreen - -NO 3 - /NH 4 + N/A + RS mRed 0.5 -NO 3 - /NH 4 + II + RS mRed 0.8 -NH 4 + V + RS mRed 0.5 -NH 4 + N/A + RS mGreen - -NH 4 + N/A + RS mGreen - -NH 4 + N/A + RS mRed 0.8 -NH 4 + IV + RS mRed 0.7 -NH 4 + I + RS mGreen - -NH 4 + N/A + RS mGreen - -NH 4 + N/A + RS mOrangeNO 3 - V - RS mOrangeNO 3 - VI -

Synechococcus genes being used for multi-locus sequence typing and community structure analysis GeneEncoding rpoC1DNA-dependent polymerase petB/D fragmentsb-type cytochrome & subunit of the photosynthetic & intergenic regioncytochrome b6/f complex, respectively rnpBribonucleoprotein enzyme (RNase P) pstSperiplasmic phosphate-binding protein 16S rDNAsmall subunit rRNA ntcAglobal N transcriptional regulator

16S rRNA petB-D rpoC1pstS Distinguishing alleles petBD intergenic region - sequence length rpoC1 pstS 5% divergence 16S rDNA 1% divergence

12 C 14 C 12 C 14 C 14 C labelling of DNA : growth of Prochlorococcus NATL1-MIT on 14 C NaHCO 3 55,000 x g o/n 40,000 x g o/n

Environmental DNA received from partners : ROSCOFF BLANESHELGOLAND Name DNA amplifiable RA x x RA faint  RA faint x RA   Name DNA amplifiable BL   BL   Name DNA amplifiable HE   HE   HE x x Full length clone libraries generated : pA (8-28) pHr ( ) Site Primers Identifier HE pA/pHrHe(H) 25/02/01 Mio (Prosope cruise) pA/pHrMio(M) 25/02/01 Dyf (Prosope cruise) pA/pHrDyf (D) 25/02/01

Prosope cruise track

Sample Name DYF MIO Helgoland Number of clones % Containing Insert % Amplified with Primer couple 359F/1313R (oxygenic-phototroph specific) % Amplified with Primer couple 359F/1038R (1038r amplifies all marine Syns & Pros, ACE/PS680 cluster, PCC9005 = marine cyano-specific) Analysis of full length environmental clone libraries

Sequence analysis of Cyanobacterial/plastid sequences from full length clone libraries SequenceClosest BLAST matchidentity DYF13Prochlorococcus ENATL4, PAC1 99% DYF33Prochlorococcus ENATL4, PAC1 99% HE24env.SAR7; Synechococcus MIO12Prochlorococcus MIT % MIO31Prochlorococcus ENATL4, PAC1 99% MIO43Prochlorococcus ENATL4, PAC1 99% MIO60Prochlorococcus MIT9211; MIT % DYF18env. Plastid seqs. OCS20; OM283 (cryptomonad) 99% DYF25env. Plastid seqs. OCS50, OM153; (Haptophyte) 97% DYF38Skeletonema pseudocostatum, uncult. Bacillariophyte 96% env. Plastid seq. Diatom DYF45Chrysochromulina polyepsis, env. Plastid OCS31; 98% HE3 HE3Chrysochromulina polyepsis, env. Plastid OCS31; 98% DYF45 HE12 env. Seq. OCS50; E. huxleyi plastid 100% HE17 env. Plastid seq. OM81; Ochromonas sp. plastid 93% MIO26 env. Plastid OCS31, OM21; 98%

Analysis of 107F-1038R environmental clone libraries Site Total no. of clones No. of unique RFLP types No. of unique clones Coverage (%) (HaeIII + EcoRI) HE HE RA RA BL000921* BL * of 9 clones partially sequenced 8 are Syns or Pros 1 has closest homology to a magnetitie containing magnetic vibrio but the identity is low (85.2%)