Quantitative dynamics of triacylglycerol accumulation in microalgae populations at single-cell resolution revealed by Raman microspectroscopy.
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.
Single-Cell Center, CAS Key Laboratory of Biofuels and Shandong Key Laboratory of Energy Genetics, Qingdao Institute of BioEnergy and Bioprocess Technology, Chinese Academy of Sciences, 189 Songling Road, Qingdao, Shandong 266101, China.
Tingting
T
Wang
Single-Cell Center, CAS Key Laboratory of Biofuels and Shandong Key Laboratory of Energy Genetics, Qingdao Institute of BioEnergy and Bioprocess Technology, Chinese Academy of Sciences, 189 Songling Road, Qingdao, Shandong 266101, China.
Yuetong
Y
Ji
Single-Cell Center, CAS Key Laboratory of Biofuels and Shandong Key Laboratory of Energy Genetics, Qingdao Institute of BioEnergy and Bioprocess Technology, Chinese Academy of Sciences, 189 Songling Road, Qingdao, Shandong 266101, China.
Yun
Y
Wang
Single-Cell Center, CAS Key Laboratory of Biofuels and Shandong Key Laboratory of Energy Genetics, Qingdao Institute of BioEnergy and Bioprocess Technology, Chinese Academy of Sciences, 189 Songling Road, Qingdao, Shandong 266101, China.
Jing
J
Jia
Single-Cell Center, CAS Key Laboratory of Biofuels and Shandong Key Laboratory of Energy Genetics, Qingdao Institute of BioEnergy and Bioprocess Technology, Chinese Academy of Sciences, 189 Songling Road, Qingdao, Shandong 266101, China.
Jing
J
Li
Single-Cell Center, CAS Key Laboratory of Biofuels and Shandong Key Laboratory of Energy Genetics, Qingdao Institute of BioEnergy and Bioprocess Technology, Chinese Academy of Sciences, 189 Songling Road, Qingdao, Shandong 266101, China.
Shi
S
Huang
Laboratory for Algae Research and Biotechnology, College of Technology and Innovation, Arizona State University, 7417 E. Unity Avenue, Mesa, Arizona 85212, USA.
Danxiang
D
Han
Laboratory for Algae Research and Biotechnology, College of Technology and Innovation, Arizona State University, 7417 E. Unity Avenue, Mesa, Arizona 85212, USA.
Qiang
Q
Hu
Kroto Research Institute, University of Sheffield, Broad Lane, Sheffield, South Yorkshire S3 7HQ, United Kingdom.
Wei E
WE
Huang
Single-Cell Center, CAS Key Laboratory of Biofuels and Shandong Key Laboratory of Energy Genetics, Qingdao Institute of BioEnergy and Bioprocess Technology, Chinese Academy of Sciences, 189 Songling Road, Qingdao, Shandong 266101, China.
Jian
J
Xu
Bioprocess dynamics; Microalgae; Population heterogeneity; Single-cell Raman spectra; Triacylglycerol