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Screening of three Chlorella mutant strains with high lipid production induced by 3 types of lasers

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Abstract

There are many studies on the use of lasers as a special mutation-breeding method to select microorganisms and plant seeds. In this study, we screened 72 microalgal mutants from the Chlorella samples irradiated using three types of laser (He-Ne, Nd:YAG and SC). Finally, 2 mutants N2 and S3 from wild Chlorella pyrenoidosa, and 1 mutant, N3 from Chlorella pacifica having high potential for biodiesel applications with high yield and stability, were obtained after comprehensive comparison and systematic screening. The average total lipid productivities of the 3 mutants were 19.90 ± 2.46 mg L−1 day−1 for N2, 29.82 ± 4.45 mg L−1 day−1 for S3 and 20.56 ± 5.69 mg L−1 day−1 for N3, respectively. The 3 mutants were proved they could display higher lipid productivity, lipid content, growth rate and transcriptional expression trends of 7 genes relevant to fatty acid biosynthesis than the wild strain in 10 continuous generation subcultures.

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References

  • Campbell WH (1988) Nitrate reductase and its role in nitrate assimilation in plants. Physiol Plant 74:214–219

    CAS  Google Scholar 

  • Chakraborty S, Mohanty D, Ghosh S, Das D (2016) Improvement of lipid content of Chlorella minutissima MCC 5 for biodiesel production. J Biosci Bioeng 122:294–300

    CAS  PubMed  Google Scholar 

  • Crawford NM, Smith M, Bellissimo D, Davis RW (1988) Sequence and nitrate regulation of the Arabidopsis thaliana mRNA encoding nitrate reductase, a metalloflavoprotein with three functional domains. Proc Natl Acad Sci 85:5006–5010

    CAS  PubMed  Google Scholar 

  • Damiani MC, Popovich CA, Constenla D, Leonardi P (2010) Lipid analysis in Haematococcus pluvialis to assess its potential use as a biodiesel feedstock. Bioresour Technol 101:3801–3807

    CAS  PubMed  Google Scholar 

  • Doan TTY, Obbard JP (2012) Enhanced intracellular lipid in Nannochloropsis sp. via random mutagenesis and flow cytometric cell sorting. Algal Research 1:17–21

  • Drobetsky EA, Turcotte J, Chateauneuf A (1995) A role for ultraviolet A in solar mutagenesis. Proc Natl Acad Sci USA. 92:2350–2354

  • Du HM, Ahmed F, Lin B, Li Z, Huang YH, Sun G, Ding H, Wang C, Meng CX, Gao ZQ (2017) The effects of plant growth regulators on cell growth, protein, carotenoid, PUFAs and lipid production of C. pyrenoidosa ZF strain. Energies 10:1696

    Google Scholar 

  • Feng Y, Li C, Zhang D (2011) Lipid production of Chlorella vulgaris cultured in artificial wastewater medium. Bioresour Technol 102:101–105

    CAS  PubMed  Google Scholar 

  • Freitas AA, Magalhães JPD (2011) A review and appraisal of the DNA damage theory of ageing. Mutat Res 728:12–22

    CAS  PubMed  Google Scholar 

  • Gao ZQ, Li DM, Meng CX, Xu D, Zhang XW, Ye NH (2013) Survival and proliferation characteristics of the microalga Chlamydomonas sp ICE-L after hypergravitational stress pretreatment. Icarus 226:971–979

    Google Scholar 

  • Greene JC, Whitworth AJ, Kuo I, Andrews LA, Feany MB, Pallanck LJ (2003) Mitochondrial pathology and apoptotic muscle degeneration in Drosophila parkin mutants. Proc Natl Acad Sci USA 100(7): 4078–83

  • Huang WP, Lin Y, He M, Gong Y, Huang J (2018) Induced high-yield production of zeaxanthin, lutein, and β-carotene by a mutant of Chlorella zofingiensis. J Agric Food Chem 66:891–897

    CAS  PubMed  Google Scholar 

  • Gao ZQ, Miao XX, Zhang XW, Wu GX, Guo YY, Wang MM, Bang L, Li XB, Gao YB, Hu S (2016) Comparative fatty acid transcriptomic test and iTRAQ-based proteomic analysis in Haematococcus pluvialis upon salicylic acid (SA) and jasmonic acid (JA) inductions. Algal Res 17:277–284

    Google Scholar 

  • Guillard RRL (1975) Culture of phytoplankton for feeding marine invertebrates. In: Smith WL, Chanley MH (eds) Culture of marine invertebrate animals. Plenum Press, New York, pp 29–60

    Google Scholar 

  • Han JR, Wang Q (2010) Laser mutagenesis of protoplasts of Penicillium sp. PT95 for the enhancement of carotenoid yield. J Appl Microbiol 105:1552–1557

    Google Scholar 

  • Han R, Wang X, Yue M (2002) Influence of He-Ne laser irradiation on the excision repair of cyclobutyl pyrimidine dimers in the wheat DNA. Chin Sci Bull 47:818–821

    CAS  Google Scholar 

  • Hlavova M, Turoczy Z, Bisova K (2015) Improving microalgae for biotechnology–from genetics to synthetic biology. Biotechnol Adv 33:1194–1203

    CAS  PubMed  Google Scholar 

  • Hu Q, Sommerfeld M, Jarvis E, Ghirardi M, Posewitz MM, Darzins A (2010) Microalgal triacylglycerols as feedstocks for biofuel production: perspectives and advances. Plant J 54:621–639

    Google Scholar 

  • Josephine A, Niveditha C, Radhika A, Shali AB, Kumar TS, Dharani G, Kirubagaran R (2015) Analytical evaluation of different carbon sources and growth stimulators on the biomass and lipid production of Chlorella vulgaris–implications for biofuels. Biomass Bioenergy 75:170–179

    CAS  Google Scholar 

  • Kao CY, Chiu SY, Huang TT, Dai L, Wang GH, Tseng CP, Chen CH, Lin CS (2012) A mutant strain of microalga Chlorella sp. for the carbon dioxide capture from biogas. Biomass Bioenergy 36:132–140

    CAS  Google Scholar 

  • Karu T, Tiphlova O, Esenaliev R, Letokhov V (1994) Two different mechanisms of low-intensity laser photobiological effects on Escherichia coli. J Photochem Photobiol B Biol 24:155–161

    CAS  Google Scholar 

  • Khozin-Goldberg I, Cohen Z (2011) Unraveling algal lipid metabolism: recent advances in gene identification. Biochimie 93:91–100

    CAS  PubMed  Google Scholar 

  • Kim DG, Lee C, Park S-M, Choi Y-E (2014) Manipulation of light wavelength at appropriate growth stage to enhance biomass productivity and fatty acid methyl ester yield using Chlorella vulgaris. Bioresour Technol 159:240-248

  • Lee B, Choi GG, Choi YE, Sung M, Park MS, Yang JW (2014) Enhancement of lipid productivity by ethyl methane sulfonate-mediated random mutagenesis and proteomic analysis in Chlamydomonas reinhardtii. Korean J Chem Eng 31:1036–1042

    CAS  Google Scholar 

  • Lei AP, Chen H, Shen GM, Hu ZL, Chen L, Wang JX (2012) Expression of fatty acid synthesis genes and fatty acid accumulation in Haematococcus pluvialis under different stressors. Biotechnol Biofuel 5:1–18

    Google Scholar 

  • Leyva LA, Bashan Y, de Bashan LE (2015) Activity of acetyl-CoA carboxylase is not directly linked to accumulation of lipids when Chlorella vulgaris is co-immobilised with Zospirillum brasilense in alginate under autotrophic and heterotrophic conditions. Ann Microbiol 65:339–349

    CAS  Google Scholar 

  • Li YT, Han DX, Sommerfeld M, Hu Q (2011) Photosynthetic carbon partitioning and lipid production in the oleaginous microalga Pseudochlorococcum sp. (Chlorophyceae) under nitrogen-limited conditions. Bioresour Technol 102:123–129

    CAS  PubMed  Google Scholar 

  • Lin B, Ahmed F, Du HM, Li Z, Yan YC, Huang YH, Cui M, Yin YH, Li B, Wang MM, Meng CX, Gao ZQ (2018) Plant growth regulators promote lipid and carotenoid accumulation in Chlorella vulgaris. J Appl Phycol 30:1549–1561

    CAS  Google Scholar 

  • Livak KJ, Schmittgen TD (2001) Analysis of relative gene expression data using real-time quantitative PCR and the 2-△△CT method. Methods 25:402–408

    CAS  Google Scholar 

  • Liu F, Chen H, Han R (2015b) The effects of He-Ne laser and enhanced ultraviolet-B radiation on proliferating-cell nuclear antigen in wheat seedlings. Amer J Plant Sci 6:1206–1214

    CAS  Google Scholar 

  • Liu S, Zhao Y, Liu L, Ao X, Ma L, Wu M, Ma F (2015a) Improving cell growth and lipid accumulation in green microalgae Chlorella sp. via UV irradiation. Appl Biochem Biotechnol 175:3507–3518

    CAS  PubMed  Google Scholar 

  • Ma YB, Wang ZY, Zhu M, Yu CJ, Cao YP, Zhang DY, Zhou GK (2013) Increased lipid productivity and TAG content in Nannochloropsis by heavy-ion irradiation mutagenesis. Bioresour Technol 136:360–367

    CAS  PubMed  Google Scholar 

  • Manandhar-Shrestha K, Hildebrand M (2013) Development of flow cytometric procedures for the efficient isolation of improved lipid accumulation mutants in a Chlorella sp. microalga. J Appl Phycol 25:1643–1651

    CAS  PubMed  PubMed Central  Google Scholar 

  • Manirakiza P, Covaci A, Schepens P (2001) Comparative study on total lipid determination using Soxhlet, Roese-Gottlieb, Bligh & Dyer, and modified Bligh & Dyer extraction methods. J Food Compos Anal 14:93–100

    CAS  Google Scholar 

  • McDermott C, Nho C, Howard W, Holton B (1998) The cyanobacterial toxin, microcystin-LR, can induce apoptosis in a variety of cell types. Toxicon 36:1981–1996

    CAS  PubMed  Google Scholar 

  • Montreuil AL, Bullard J, Chandler J (2013) Detecting seasonal variations in embryo dune morphology using a terrestrial laser scanner. J Coast Res 118:1313–1318

    Google Scholar 

  • Najafi MBH (2013) Bacterial mutation; types, mechanisms and mutant detection methods: a review. Eur Sci J 4:1857–7431

    Google Scholar 

  • Ong SC, Kao CY, Chiu SY, Tsai MT, Lin CS (2010) Characterization of the thermaltolerant mutants of Chlorella sp. with high growth rate and application in outdoor photobioreactor cultivation. Bioresour Technol 101:2880–2883

    CAS  PubMed  Google Scholar 

  • Perez-Garcia O, Escalante FME, Bashan LE, Bashan Y (2011). Heterotrophic cultures of microalgae: metabolism and potential products. Water Res 45:11–36

  • Přibyl P, Cepák V, Zachleder V (2012) Production of lipids in 10 strains of Chlorella and Parachlorella, and enhanced lipid productivity in Chlorella vulgaris. Appl Microbiol Biotechnol 94:549–561

    PubMed  Google Scholar 

  • Riekhof WR, Sears BB, Benning C (2005) Annotation of genes involved in glycerol lipid biosynthesis in Chlamydomonas reinhardtii: discovery of the betaine lipid synthase BTA1Cr. Eukaryot Cell 4:242–252

    CAS  PubMed  PubMed Central  Google Scholar 

  • Rhodes L, Smith J, Tervit R, Roberts R, Adamson J, Adams S, Decker M (2006) Cryopreservation of economically valuable marine micro-algae in the classes Bacillariophyceae, Chlorophyceae, Cyanophyceae, Dinophyceae, Haptophyceae, Prasinophyceae, and Rhodophyceae. Cryobiol 52:152–156

    CAS  Google Scholar 

  • Rosenberg JN, Oyler GA, Wilkinson L, Betenbaugh MJ (2008) A green light for engineered algae: redirecting metabolism to fuel a biotechnology revolution. Curr Opin Biotechnol 19:430–436

    CAS  PubMed  Google Scholar 

  • Sachdeva N, Gupta RP, Mathur AS, Tuli DK (2016) Enhanced lipid production in thermo-tolerant mutants of Chlorella pyrenoidosa NCIM 2738. Bioresour Technol 221:576–587

    CAS  PubMed  Google Scholar 

  • Salie MJ, Thelen JJ (2016) Regulation and structure of the heteromeric acetyl-CoA carboxylase. Biochim Biophys Acta-Mol Cell Biol Lip 1861:1207–1213

    CAS  Google Scholar 

  • Sarayloo E, Simsek S, Unlu YS, Cevahir G, Erkey C, Kavakli IH (2018) Enhancement of the lipid productivity and fatty acid methyl ester profile of Chlorella vulgaris by two rounds of mutagenesis. Bioresour Technol 250:764–769

    CAS  PubMed  Google Scholar 

  • Shekh AY, Shrivastava P, Krishnamurthi K, Mudliar SN, Devi SS, Kanade GS, Chakrabarti T (2016) Stress enhances poly-unsaturation rich lipid accumulation in Chlorella sp. and Chlamydomonas sp. Biomass Bioenergy 84:59–66

    CAS  Google Scholar 

  • Singh DK, Kumar S (2008) Nitrate reductase, arginine deaminase urease and dehydrogenase activities in natural soil (ridges with forest) and in cotton soil after acetamiprid treatments. Chemosphere 71:412–418

    CAS  PubMed  Google Scholar 

  • Sivaramakrishnan R, Incharoensakdi A (2017) Enhancement of lipid production in Scenedesmus sp. by UV mutagenesis and hydrogen peroxide treatment. Bioresour Technol 235:366–370

  • Tanadul O, Noochanong W, Jirakranwong P, Chanprame S (2018) EMS-induced mutation followed by quizalofop-screening increased lipid productivity in Chlorella sp. Bioprocess Biosyst Eng 41:613

    CAS  PubMed  Google Scholar 

  • Wang LQ, Yang GZ, Chen FD (1997) Biological effects and application in plants and animals heredity and breeding by laser mutation. Acta Laser Biol Sin 1997:1097–1102

    Google Scholar 

  • Wang WL, Wei TT, Fan JH, Yi J, Li YG, Wan MX, Wang J, Bai WM (2018) Repeated mutagenic effects of 60Co-γ irradiation coupled with high throughput screening improves lipid accumulation in mutant strains of the microalgae Chlorella pyrenoidosa as a feedstock for bioenergy. Algal Res 33:71–77

    Google Scholar 

  • Xie B, Stessman D, Hart JH, Dong HL, Wang YJ, Wright DA, Nikolau BJ, Spalding MH, Halverson LJ (2014) High-throughput fluorescence-activated cell sorting for lipid hyperaccumulating Chlamydomonas reinhardtii mutants. Plant Biotechnol 12:872–882

    CAS  Google Scholar 

  • Zabaglo L, Ormerod MG, Dowsett M (2015) Measurement of markers for breast cancer in a model system using laser scanning cytometry. Cytometry 41:166–171

    Google Scholar 

  • Zhang HN, Gao ZQ, Li Z, Du HM, Lin B, Cui M, Yin YH, Lei FM, Yu CY, Meng CX (2017) Laser radiation induces growth and lipid accumulation in the seawater microalga Chlorella pacifica. Energies 10:1671

    Google Scholar 

  • Zhao YS, Wang LY, Zheng H, Yin HP, Chen XD, Tan ZT, Wu WT (1999) Effect of frequency-doubling pulse Nd:YAG laser on microbial mutation. Proc. SPIE 3863, 1999 International Conference on Biomedical Optics

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Funding

The present study was supported by National Natural Science Foundation of China (31972815), Major Basic Research Program for Natural Science Foundation of Shandong Province (ZR2019ZD17), Key Research and Development Program of Shandong Province (Food for Special Medical Purpose) (2018YYSP016), Key Research and Development Program of Shandong Province (2017CXGC0309) and the Project of Shandong Province Higher Educational Science and Technology Program (J17KA132).

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In this work, Chunxiao Meng and Zhengquan Gao had completed the experimental design. Kang Wang and Bin Lin had completed data analysis, thesis writing, RNA isolation and real-time qPCR. Zhe Li had completed laser mutagenesis. Haonan Zhang, Huanmin Du and Yanyun Guo had completed microalgal screening, culture and lipid content detection. Fei Xu and Xiao Jiang had completed enzyme activity assay.

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Correspondence to Chunxiao Meng or Zhengquan Gao.

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Bin Lin and Kang Wang are co-first authors

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Wang, K., Lin, B., Meng, C. et al. Screening of three Chlorella mutant strains with high lipid production induced by 3 types of lasers. J Appl Phycol 32, 1655–1668 (2020). https://doi.org/10.1007/s10811-020-02112-5

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