Cite this paper:
CAO Min, ZHANG Fangfang, MAO Yunxiang, KONG Fanna, WANG Dongmei. Characterization of the squalene-rich Botryococcus braunii Abt02 strain[J]. HaiyangYuHuZhao, 2019, 37(2): 675-684

Characterization of the squalene-rich Botryococcus braunii Abt02 strain

CAO Min1,2, ZHANG Fangfang1,2, MAO Yunxiang1,2,3, KONG Fanna1,2, WANG Dongmei1,2
1 College of Marine Life Sciences, Ocean University of China, Qingdao 266003, China;
2 Key Laboratory of Marine Genetics and Breeding(OUC), Ministry of Education, Qingdao 266003, China;
3 Laboratory for Marine Biology and Biotechnology, Qingdao National Laboratory for Marine Science and Technology, Qingdao 266000, China
Abstract:
Botryococcus braunii is widely studied due to its high hydrocarbon content. In this study, B. brauniiAbt02 was subjected to several analyses, including cytological observation, hydrocarbon composition analysis by gas chromatography mass spectrometry (GC-MS), phylogenetic identification using known races (A, B and L) of B. braunii strains based on their 18S rDNA sequences, and qPCR-based investigation of transcript accumulation levels of hydrocarbon biosynthesis-related enzymes (DXS, MCS, DLS, SQS) during different growth phases (lag phase, log phase, early stationary growth phase, late stationary growth phase) under nitrogen-replete and nitrogen-depleted growth conditions, respectively. Based on cytological observation and on the 18S rDNA phylogenetic analysis, strain Abt02 was assigned to race B. Analysis of the strain's chemical composition showed that the B. braunii Abt02 contained high levels of hydrocarbons, which accounted for 43.75% of the cell's dry weight. Of these hydrocarbons, squalene and its derivatives accounted for up to 87.54%. In addition, all four enzymes investigated were expressed at higher levels during the log growth phase under nitrogen depleted conditions than under nitrogen replete conditions.
Key words:    Botryococcus braunii|cytological observation|phylogenetic analysis|hydrocarbon components|qPCR   
Received: 2018-03-13   Revised: 2018-05-15
Tools
PDF (899 KB) Free
Print this page
Add to favorites
Email this article to others
Authors
Articles by CAO Min
Articles by ZHANG Fangfang
Articles by MAO Yunxiang
Articles by KONG Fanna
Articles by WANG Dongmei
References:
Banerjee A, Sharma R, Chisti Y, Banerjee U C. 2002. Botryococcus braunii:a renewable source of hydrocarbons and other chemicals. Critical Reviews in Biotechnology, 22(3):245-279.
Brown A C, Knights B A, Conway E. 1969. Hydrocarbon content and its relationship to physiological state in the green alga Botryococcus braunii. Phytochemistry, 8(3):543-547.
Chiang I Z, Huang W Y, Wu J T. 2004. Allelochemicals of Botryococcus braunii (Chlorophyceae). Journal of Phycology, 40(3):474-480.
Choi G G, Kim B H, Ahn C Y, Oh H M. 2011. Effect of nitrogen limitation on oleic acid biosynthesis in Botryococcus braunii. Journal of Applied Phycology, 23(6):1 031-1 037.
Fang L, Sun D Y, Xu Z Y, He J, Qi S Y, Chen X, Chew W, Liu J H. 2015. Transcriptomic analysis of a moderately growing subisolate Botryococcus braunii 779(Chlorophyta) in response to nitrogen deprivation.Biotechnology for Biofuels, 8:130.
Fernandes N V, Yeganehjoo H, Katuru R, DeBose-Boyd R A, Morris L L, Michon R, Yu Z L, Mo H B. 2013. Geranylgeraniol suppresses the viability of human DU145 prostate carcinoma cells and the level of HMG CoA reductase. Experimental Biology and Medicine, 238(11):1 265-1 274.
Hillen L W, Pollard G, Wake L V, White N. 1982. Hydrocracking of the oils of Botryococcus braunii to transport fuels.Biotechnology and Bioengineering, 24(1):193-205.
Huss V A R, Sogin M L. 1990. Phylogenetic position of some Chlorella species within the chlorococcales based upon complete small-subunit ribosomal RNA sequences.Journal of Molecular Evolution, 31(5):432-442.
Ioki M, Baba M, Bidadi H, Suzuki I, Shiraiwa Y, Watanabe M M, Nakajima N. 2012. Modes of hydrocarbon oil biosynthesis revealed by comparative gene expression analysis for race A and race B strains of Botryococcus braunii. Bioresource Technology, 109:271-276.
Jarstfer M B, Blagg B S J, Rogers D H, Poulter C D. 1996. Biosynthesis of squalene. Evidence for a tertiary cyclopropylcarbinyl cationic intermediate in the rearrangement of presqualene diphosphate to squalene.Journal of the American Chemical Society, 118(51):13 089-13 090.
Kawachi M, Tanoi T, Demura M, Kaya K, Watanabe M M. 2012. Relationship between hydrocarbons and molecular phylogeny of Botryococcus braunii. Algal Research, 1(2):114-119.
Larkin M A, Blackshields G, Brown N P, Chenna R, McGettigan P A, McWilliam H, Valentin F, Wallace I M, Wilm A, Lopez P, Thompson J D, Gibson T J, Higgins D G. 2007. Clustal W and Clustal X version 2.0. Bioinformatics, 23(21):2 947-2 948.
Liao Z H, Chen M, Gong Y F, Miao Z Q, Sun X F, Tang K X. 2006. Isoprenoid biosynthesis in plants:pathways, genes, regulation and metabolic engineering. Journal of Biological Sciences, 6(1):209-219.
Liu X Y. 2013. Studies on the Separation, Identification and Mutation Breeding of Botryococcus braunii. Zhejiang University, Hangzhou. (in Chinese)
Matsushima D, Jenke-Kodama H, Sato Y, Fukunaga Y, Sumimoto K, Kuzuyama T, Matsunaga S, Okada S. 2012. The single cellular green microalga Botryococcus braunii, race B possesses three distinct 1-deoxy-D-xylulose 5-phosphate synthases. Plant Science, 185-186:309-320.
Metzger P, Allard B, Casadevall E, Berkaloff C, Couté A. 1990. Structure and chemistry of a new chemical race of Botryococcus braunii (Chlorophyceae) that produces lycopadiene, a tetraterpenoid hydrocarbon. Journal of Phycology, 26(2):258-266.
Metzger P, Berkaloff C, Casadevall E, Coute A. 1985. Alkadiene-and botryococcene-producing races of wild strains of Botryococcus braunii. Phytochemistry, 24(10):2 305-2 312.
Metzger P, Casadevall E, Coute A. 1988. Botryococcene distribution in strains of the green alga Botryococcus braunii. Phytochemistry, 27(5):1 383-1 388.
Metzger P, Casadevall E. 1989. Aldehydes, very long chain alkenylphenols, epoxides and other lipids from an alkadiene-producing strain of Botryococcus braunii.Phytochemistry, 28(8):2 097-2 104.
Metzger P, Largeau C. 2005. Botryococcus braunii:a rich source for hydrocarbons and related ether lipids. Applied Microbiology and Biotechnology, 66(5):486-496.
Niehaus T D, Okada S, Devarenne T P, Watt D S, Sviripa V, Chappell J. 2011. Identification of unique mechanisms for triterpene biosynthesis in Botryococcus braunii.Proceedings of the National Academy of Sciences of the United States of America, 108(30):12 260-12 265.
Poulter C D. 1990. Biosynthesis of non-head-to-tail terpenes.Formation of 1'-1 and 1'-3 linkages. Accounts of Chemical Research, 23(3):70-77.
Sato Y, Ito Y, Okada S, Murakami M, Abe H. 2003. Biosynthesis of the triterpenoids, botryococcenes and tetramethylsqualene in the B race of Botryococcus braunii via the non-mevalonate pathway. Tetrahedron Letters, 44(37):7 035-7 037.
Schwender J, Seemann M, Lichtenthaler H K, Rohmer M. 1996. Biosynthesis of isoprenoids (carotenoids, sterols, prenyl side-chains of chlorophylls and plastoquinone) via a novel pyruvate/glyceraldehyde 3-phosphate nonmevalonate pathway in the green alga Scenedesmus obliquus. Biochemical Journal, 316(Pt 1):73-80.
Senousy H H, Beakes G W, Hack E. 2004. Phylogenetic placement of Botryococcus braunii (Trebouxiophyceae) and Botryococcus sudeticus isolate UTEX 2629(Chlorophyceae). Journal of Phycology, 40(2):412-423.
Singh Y, Kumar H D. 1992. Lipid and hydrocarbon production by Botryococcus spp. under nitrogen limitation and anaerobiosis. World Journal of Microbiology and Biotechnology, 8(2):121-124.
Talukdar J, Kalita M C, Goswami B C. 2013. Characterization of the biofuel potential of a newly isolated strain of the microalga Botryococcus braunii Kützing from Assam, India. Bioresource Technology, 149:268-275.
Tamura K, Stecher G, Peterson D, Filipski A, Kumar S. 2013. MEGA6:molecular evolutionary genetics analysis version 6.0. Molecular Biology and Evolution, 30(12):2 725-2 729.
Tanoi T, Kawachi M, Watanabe M M. 2011. Effects of carbon source on growth and morphology of Botryococcus braunii. Journal of Applied Phycology, 23(1):25-33.
Wake L V, Hillen L W. 1980. Study of a "bloom" of the oil-rich alga Botryococcus braunii in the Darwin River Reservoir.Biotechnology and Bioengineering, 22(8):1 637-1 656.
Wake L V, Hillen L W. 1981. Nature and hydrocarbon content of blooms of the alga Botryococcus braunii occuring in Australian freshwater lakes. Marine and Freshwater Research, 32(3):353-367.
Wang P Y, Mao Y X, Kong F N, Ma M, Ma F. 2011. Morphological and genetic diversity of Botryococcus braunii. Periodical of Ocean University of China, 41(5):63-70. (in Chinese)
White J D, Somers T C, Reddy G N. 1986. The absolute configuration of (-)-botryococcene. Journal of the American Chemical Society, 108(17):5 352-5 353.
White J D, Somers T C, Reddy G N. 1992. Degradation and absolute configurational assignment to C34-botryococcene. The Journal of Organic Chemistry, 57(18):4 991-4 998.
Zhang D L, Poulter C D. 1995. Biosynthesis of non-head-totail isoprenoids. Synthesis of 1'-1 and 1'-3 structures by recombinant yeast squalene synthase. Journal of the American Chemical Society, 117(5):1 641-1 642.