Metabolic phenotyping of the cyanobacterium Synechocystis 6803 engineered for production of alkanes and free fatty acids

Hu, Ping; Borglin, Sharon; Kamennaya, Nina A.; Chen, Liang; Park, Hanwool; Mahoney, Laura; Kijac, Aleksandra; Shan, George; Chavarría, Krystle L.; Zhang, Chunmin; Quinn, Nigel W.T.; Wemmer, David E.; Holman, Hoi‐Ying N.; Jansson, Christer

doi:10.1016/j.apenergy.2012.08.047

Cited by 43 publications

(29 citation statements)

References 39 publications

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“…PCC 6803 is consistent with the previous reports of Tan et al [29] and Guan et al [40]. However, these results contrast with a recent report that this strain produces 9-heptadecene [18]. Moreover, the position of the methyl group in the aliphatic chain was confirmed using the NIST mass spectral library and the analysis of the Cn-1 hydrocarbon biosynthesis mechanism.…”

Section: Discussioncontrasting

confidence: 56%

“…Although the baeocystous type (subsection II) and the ramified type (subsection V) exhibit unicellular and filamentous phenotypes, respectively [6], further study of these subsections is necessary for a thorough understanding of the hydrocarbon distribution in cyanobacteria on the phylum level. Nostoc spongiaeforme FACHB 130, which showed a greater hydrocarbon production than all wild type strains previously reported [18,29], was identified as a prospective strain for the production of hydrocarbons with a biomass productivity of 1.3 g/L DCW and a hydrocarbon yield of 1.8 mg/g DCW ( Table 2). It is well known that hydrocarbons can constitute up to 75% of the DCW of Botryococcus braunii, an eukaryotic microalga belonging to the green algae division of chlorophyta [35].…”

Section: Discussionmentioning

confidence: 99%

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Hydrocarbon profiles and phylogenetic analyses of diversified cyanobacterial species

Liu¹,

Zhu²,

Lü³

et al. 2013

Applied Energy

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Section: Discussioncontrasting

confidence: 56%

Section: Discussionmentioning

confidence: 99%

Hydrocarbon profiles and phylogenetic analyses of diversified cyanobacterial species

Liu¹,

Zhu²,

Lü³

et al. 2013

Applied Energy

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“…Two regions of the spectra: the biochemical fingerprint region (800 cm −1 to 1800 cm −1 ) and the hydrocarbon region (2600 cm −1 to 3100 cm −1 ), were extracted for further analyses in order to extract useful information contained in Raman bands from the useless noise [28,43]. For both regions a spectrum was normalized via division by its area.…”

Section: Methodsmentioning

confidence: 99%

Quantitative dynamics of triacylglycerol accumulation in microalgae populations at single-cell resolution revealed by Raman microspectroscopy

Wang

et al. 2014

Biotechnol Biofuels

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BackgroundRapid, real-time and label-free measurement of the cellular contents of biofuel molecules such as triacylglycerol (TAG) in populations at single-cell resolution are important for bioprocess control and understanding of the population heterogeneity. Raman microspectroscopy can directly detect the changes of metabolite profile in a cell and thus can potentially serve these purposes.ResultsSingle-cell Raman spectra (SCRS) of the unicellular oleaginous microalgae Nannochloropsis oceanica from the cultures under nitrogen depletion (TAG-producing condition) and nitrogen repletion (non-TAG-producing condition) were sampled at eight time points during the first 96 hours upon the onset of nitrogen depletion. Single N. oceanica cells were captured by a 532-nm laser and the SCRS were acquired by the same laser within one second per cell. Using chemometric methods, the SCRS were able to discriminate cells between nitrogen-replete and nitrogen-depleted conditions at as early as 6 hours with >93.3% accuracy, and among the eight time points under nitrogen depletion with >90.4% accuracy. Quantitative prediction of TAG content in single cells was achieved and validated via SCRS and liquid chromatography-mass spectrometry (LC-MS) analysis at population level. SCRS revealed the dynamics of heterogeneity in TAG production among cells in each isogenic population. A significant negative correlation between TAG content and lipid unsaturation degree in individual microalgae cells was observed.ConclusionsOur results show that SCRS can serve as a label-free and non-invasive proxy for quantitatively tracking and screening cellular TAG content in real-time at single-cell level. Phenotypic comparison of single cells via SCRS should also help investigating the mechanisms of functional heterogeneity within a cellular population.

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“…strain PCC7942 was designed to terminate with a multiple cloning site and was flanked with the psbAI fragments. Insertion of this construct into the pUC56-simple (Hu et al, 2013) generated the pFUEL70 plasmid ( Supplementary Fig. S1).…”

Section: Plasmids and Gene Constructsmentioning

confidence: 99%

“…Ten ml of cells in saline solution (0.9% w/v NaCl) was placed on Si-wafer chips and all the free-flowing solution was removed. All SR-FTIR absorption spectra were collected in transmission mode at a spectral resolution of 4 cm À 1 with 16 co-added scans and a peak position accuracy of 1/100 cm À 1 , which were followed by baseline removal, vector normalization, and multivariate principal componentlinear discriminant analysis (PC-LDA) using MathLab (7.0) as described in Hu et al (2013). Spectral interpretation was guided by literature (Tipson, 1968;Cabaniss and McVey, 1995;Brandenburg and Seydel, 1996;Kacȗráková et al, 2000;Naumann, 2000).…”

Section: Infrared Spectromicroscopymentioning

confidence: 99%

Installing extra bicarbonate transporters in the cyanobacterium Synechocystis sp. PCC6803 enhances biomass production

Kamennaya

Ahn

Park

et al. 2015

Metabolic Engineering

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As a means to improve carbon uptake in the cyanobacterium Synechocystis sp. strain PCC6803, we engineered strains to contain additional inducible copies of the endogenous bicarbonate transporter BicA, an essential component of the CO2-concentrating mechanism in cyanobacteria. When cultured under atmospheric CO2 pressure, the strain expressing extra BicA transporters (BicA(+) strain) grew almost twice as fast and accumulated almost twice as much biomass as the control strain. When enriched with 0.5% or 5% CO2, the BicA(+) strain grew slower than the control but still showed a superior biomass production. Introducing a point mutation in the large C-terminal cytosolic domain of the inserted BicA, at a site implicated in allosteric regulation of transport activity, resulted in a strain (BicA(+)(T485G) strain) that exhibited pronounced cell aggregation and failed to grow at 5% CO2. However, the bicarbonate uptake capacity of the induced BicA(+)(T485G) was twice higher than for the wild-type strain. Metabolic analyses, including phenotyping by synchrotron-radiation Fourier transform Infrared spectromicroscopy, scanning electron microscopy, and lectin staining, suggest that the excess assimilated carbon in BicA(+) and BicA(+)(T485G) cells was directed into production of saccharide-rich exopolymeric substances. We propose that the increased capacity for CO2 uptake in the BicA(+) strain can be capitalized on by re-directing carbon flux from exopolymeric substances to other end products such as fuels or high-value chemicals.

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Metabolic phenotyping of the cyanobacterium Synechocystis 6803 engineered for production of alkanes and free fatty acids

Cited by 43 publications

References 39 publications

Hydrocarbon profiles and phylogenetic analyses of diversified cyanobacterial species

Hydrocarbon profiles and phylogenetic analyses of diversified cyanobacterial species

Quantitative dynamics of triacylglycerol accumulation in microalgae populations at single-cell resolution revealed by Raman microspectroscopy

Installing extra bicarbonate transporters in the cyanobacterium Synechocystis sp. PCC6803 enhances biomass production

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