Table 1

Sequences of new hydrazine synthase primers targeting the hzsA subunit of anammox bacteriaa

PrimerbPosition (bp)Sequence
Forward
    hzsA_370F370–3895′-CACAAGAADGGYGGYGGDTG-3′
    hzsA_382F382–4005′-GGYGGDTGYCAGATATGGG-3′
    hzsA_526F526–5435′-TAYTTTGAAGGDGACTGG-3
    hzsA_757F757–7735′-AGTTCNAAYTWTGATCC-3′
    hzsA_757F Scalindua757–7735′-AGTTCNAAYTWTGACCC-3′
    hzsA_862F862–8785′-GTDGAYAACTGGGAYGG-3′
    hzsA_1318F1318–13385′-TATCAGCCRTTTGAYCAGGTG-3′
    hzsA_1597F1597–16155′-WTYGGKTATCARTATGTAG-3′
    hzsA_1600F1600–16175′-GGKTATCARTATGTAGAG-3′
    hzsA_1600F Scalindua1600–16205′-GGKTATCARTATGTAGAAG-3′
Reverse
    hzsA_887R868–8875′-GGATAHGCRCCRTCCCAGTT-3′
    hzsA_1353R1333–13535′-ATAHCCTTCMACHTTCACCTG-3′
    hzsA_1829R1813–18295′-TCATACCACCARTTGTA-3′
    hzsA_1829R Scalindua1813–18295′-CTGAACCACCARTTGTA-3′
    hzsA_1857R1838–18575′-AAABGGYGAATCATARTGGC-3′
    hzsA_2210R2191–22105′-GGRTTDACATAYTTWCCRCC-3
    hzsA_2390R2371–23905′-ATRTTRTCCCAYTGYGCHCC-3′
  • a The positions indicated refer to the “Ca. Kuenenia stuttgartiensis” hzsA gene (kuste2861).

  • b Several primer set were tested, but generally the primer set hzsA_526F/hzsA_1857R performed best for most of the samples. In addition, for the Ooijpolder freshwater ditches, a nested PCR with the primer set hzsA_382F/hzsA_2390R followed by the primer set hzsA_526F/hzsA_1857R or a direct PCR with the primer set hzsA_526F/hzsA_1829R was used. For the RT-PCR and qPCR, primer set hzsA_1597F/hzsA1857R is recommended. PCR products of the expected size have been obtained for Northeast Greenland and from the Barents Sea samples, using the primer combination hzsA_526F/hzsA_1829R. The primers with “Scalindua” added to their names were designed specifically to target marine “Ca. Scalindua” species.