Nitrogen fixation: Nitrogenase genes and gene expression

Zehr, Jonathan P.; Turner, Patricia J.

doi:10.1016/s0580-9517(01)30049-1

Cited by 189 publications

(185 citation statements)

References 17 publications

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“…In total, 10 ng of DNA or cDNA corresponding to 4 ng RNA was added to Pure Taq Ready-To-Go PCR Beads (GE Healthcare) along with the nifH3 and nifH4 primers in a nested PCR approach (Zehr and Turner, 2001). Illumina indices (Supplementary Table S2) were added to amplicons in the second PCR round, which were done in triplicates for each sample.…”

Section: Sampling and Environmental Parametersmentioning

confidence: 99%

Significant N2 fixation by heterotrophs, photoheterotrophs and heterocystous cyanobacteria in two temperate estuaries

et al. 2014

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Nitrogen (N) fixation is fueling planktonic production in a multitude of aquatic environments. In meso-and poly-haline estuaries, however, the contribution of N by pelagic N 2 fixation is believed to be insignificant due to the high input of N from land and the presumed absence of active N 2 -fixing organisms. Here we report N 2 fixation rates, nifH gene composition and nifH gene transcript abundance for key diazotrophic groups over 1 year in two contrasting, temperate, estuarine systems: Roskilde Fjord (RF) and the Great Belt (GB) strait. Annual pelagic N 2 fixation rates averaged 17 and 61 mmol N m À 2 per year at the two sites, respectively. In RF, N 2 fixation was mainly accompanied by transcripts related to heterotrophic (for example, Pseudomonas sp.) and photoheterotrophic bacteria (for example, unicellular diazotrophic cyanobacteria group A). In the GB, the first of two N 2 fixation peaks coincided with a similar nifH-expressing community as in RF, whereas the second peak was synchronous with increased nifH expression by an array of diazotrophs, including heterotrophic organisms as well as the heterocystous cyanobacterium Anabaena. Thus, we show for the first time that significant planktonic N 2 fixation takes place in mesohaline, temperate estuaries and that the importance of heterotrophic, photoheterotrophic and photosynthetic diazotrophs is clearly variable in space and time.

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Section: Sampling and Environmental Parametersmentioning

confidence: 99%

Significant N2 fixation by heterotrophs, photoheterotrophs and heterocystous cyanobacteria in two temperate estuaries

et al. 2014

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“…PCR amplification and 454-pyrosequencing A total of 16 DNA and 4 cDNA samples were amplified with nifH3 and nifH4 primers (Zehr and Turner, 2001) followed by 30 cycles with custom bar-coded nifH1 and nifH2 primers (Supplementary Table S1) using Pure Taq Ready-To-Go PCR Beads (GE Healthcare). As the cDNA samples from 41, 132 and 233 m yielded faint PCR products they were first amplified with nifH1 and nifH2 primers (Zehr and McReynolds, 1989) followed by 10 PCR cycles with barcoded primers (Farnelid et al, 2011).…”

Section: Environmental Samplingmentioning

confidence: 99%

Active nitrogen-fixing heterotrophic bacteria at and below the chemocline of the central Baltic Sea

et al. 2013

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The Baltic Sea receives large nitrogen inputs by diazotrophic (N 2 -fixing) heterocystous cyanobacteria but the significance of heterotrophic N 2 fixation has not been studied. Here, the diversity, abundance and transcription of the nifH fragment of the nitrogenase enzyme in two basins of the Baltic Sea proper was examined. N 2 fixation was measured at the surface (5 m) and in anoxic water (200 m). Vertical sampling profiles of 410 and o10 lm size fractions were collected in 2007, 2008 and 2011 at the Gotland Deep and in 2011 in the Bornholm Basin. Both of these stations are characterized by permanently anoxic bottom water. The 454-pyrosequencing nifH analysis revealed a diverse assemblage of nifH genes related to alpha-, beta-and gammaproteobacteria (nifH cluster I) and anaerobic bacteria (nifH cluster III) at and below the chemocline. Abundances of genes and transcripts of seven diazotrophic phylotypes were investigated using quantitative polymerase chain reaction revealing abundances of heterotrophic nifH phylotypes of up to 2.1 Â 10 7 nifH copies l À 1 . Abundant nifH transcripts (up to 3.2 Â 10 4 transcripts l À 1 ) within nifH cluster III and co-occurring N 2 fixation (0.44 ± 0.26 nmol l À 1 day À 1 ) in deep water suggests that heterotrophic diazotrophs are fixing N 2 in anoxic ammonium-rich waters. Our results reveal that N 2 fixation in the Baltic Sea is not limited to illuminated N-deplete surface waters and suggest that N 2 fixation could also be of importance in other suboxic regions of the world's oceans. The ISME Journal (2013Journal ( ) 7, 1413Journal ( -1423 doi:10.1038/ismej.2013 published online 28 February 2013 Subject Category: microbial ecology and functional diversity of natural habitats

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“…nifH PCR amplification and amplicon pyrosequencing A nested PCR protocol was used to amplify an~359-bp region of the nitrogenase gene, using the degenerate primers: nifH3, nifH4, nifH1, and nifH2 (Zani et al, 2000;Zehr and Turner, 2001). Equal volumes of DNA or cDNA were used as template (2 μl) in the first stage of the reaction, and 1 μl of PCR product was used as template in the second stage, using previously described reaction conditions (Messer et al, 2015;see Supplementary Information).…”

Section: Nucleic Acid Collection and Extractionmentioning

confidence: 99%

High levels of heterogeneity in diazotroph diversity and activity within a putative hotspot for marine nitrogen fixation

et al. 2015

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Australia's tropical waters represent predicted 'hotspots' for nitrogen (N 2 ) fixation based on empirical and modelled data. However, the identity, activity and ecology of diazotrophs within this region are virtually unknown. By coupling DNA and cDNA sequencing of nitrogenase genes (nifH) with sizefractionated N 2 fixation rate measurements, we elucidated diazotroph dynamics across the shelf region of the Arafura and Timor Seas (ATS) and oceanic Coral Sea during Austral spring and winter. During spring, Trichodesmium dominated ATS assemblages, comprising 60% of nifH DNA sequences, while Candidatus Atelocyanobacterium thalassa (UCYN-A) comprised 42% in the Coral Sea. In contrast, during winter the relative abundance of heterotrophic unicellular diazotrophs (δ-proteobacteria and γ-24774A11) increased in both regions, concomitant with a marked decline in UCYN-A sequences, whereby this clade effectively disappeared in the Coral Sea. Conservative estimates of N 2 fixation rates ranged from o1 to 91 nmol l − 1 day − 1 , and size fractionation indicated that unicellular organisms dominated N 2 fixation during both spring and winter, but average unicellular rates were up to 10-fold higher in winter than in spring. Relative abundances of UCYN-A1 and γ-24774A11 nifH transcripts negatively correlated to silicate and phosphate, suggesting an affinity for oligotrophy. Our results indicate that Australia's tropical waters are indeed hotspots for N 2 fixation and that regional physicochemical characteristics drive differential contributions of cyanobacterial and heterotrophic phylotypes to N 2 fixation.

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Nitrogen fixation: Nitrogenase genes and gene expression

Cited by 189 publications

References 17 publications

Significant N2 fixation by heterotrophs, photoheterotrophs and heterocystous cyanobacteria in two temperate estuaries

Significant N2 fixation by heterotrophs, photoheterotrophs and heterocystous cyanobacteria in two temperate estuaries

Active nitrogen-fixing heterotrophic bacteria at and below the chemocline of the central Baltic Sea

High levels of heterogeneity in diazotroph diversity and activity within a putative hotspot for marine nitrogen fixation

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