Journal
BIOTECHNOLOGY FOR BIOFUELS
Volume 7, Issue -, Pages -Publisher
BMC
DOI: 10.1186/1754-6834-7-58
Keywords
Microalgae; Triacylglycerol; Single-cell Raman spectra; Bioprocess dynamics; Population heterogeneity
Funding
- National Basic Research Program [2012CB721101]
- High-Tech Development Program [2012AA02A707]
- Methodology Innovation Program [2011IM030100]
- Ministry of Science and Technology of China
- Microevolution Program from the National Natural Science Foundation of China [91231205]
- International Innovation Partnership Program from the Chinese Academy of Sciences
- Engineering and Physical Sciences Research Council [EP/H04986X/1] Funding Source: researchfish
- EPSRC [EP/H04986X/1] Funding Source: UKRI
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Background: Rapid, 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. Results: Single-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. Conclusions: Our 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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