Cite this paper:
Fan GAO, Fangru NAN, Jia FENG, Junping LÜ, Qi LIU, Xudong LIU, Shulian XIE. Transcriptome profile of Dunaliella salina in Yuncheng Salt Lake reveals salt-stress-related genes under different salinity stresses[J]. Journal of Oceanology and Limnology, 2021, 39(6): 2336-2362

Transcriptome profile of Dunaliella salina in Yuncheng Salt Lake reveals salt-stress-related genes under different salinity stresses

Fan GAO, Fangru NAN, Jia FENG, Junping LÜ, Qi LIU, Xudong LIU, Shulian XIE
School of Life Science, Shanxi Key Laboratory for Research and Development of Regional Plants, Shanxi University, Taiyuan 030006, China
Salt stress is an abiotic stress to plants in especially saline lakes. Dunaliella, a halophilic microalga distributed throughout salt lakes and seas, can respond to different salinity stresses by regulating the expression of some genes. However, these genes and their function and biological processes involved remain unclear. Profiling these salt-stress-related genes in a high-salt-tolerant Dunaliella species will help clarify the salt tolerance machinery of Dunaliella. Three D. salina_YC salt-stress groups were tested under low (0.51 mol/L), moderate (1.03 mol/L), and high (3.42 mol/L) NaCl concentrations and one control group under very low (0.05 mol/L) NaCl concentration and 3 transcriptome results that were deep sequenced and de novo assembled were obtained per group. Twelve high-quality RNA-seq libraries with 46 585 upregulated and 47 805 downregulated unigenes were found. Relative to the control, 188 common differentially expressed genes (DEGs) were screened and divided into four clusters in expression pattern. Fifteen of them annotated in the significant enriched Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) were validated via qPCR. Their qPCR-based relative expression patterns were similar to their RNA-seq-based patterns. Two significant DEGs, the geranylgeranyl diphosphate synthase coding gene (1 876-bp cDNA) and diacylglycerol O-acyltransferase coding gene (2 968-bp cDNA), were cloned and analyzed in silico. The total lipid content, superoxide dismutase specific activity, and betacarotene content of D. salina_YC increased gradually with increasing salinity. In addition, the expression of 11 validated genes involved in fatty acid biosynthesis/degradation, active oxygen or carotenoid metabolisms showed significant changes. In addition, algal photochemical efficiency was diminished with increasing salinity, as well as the expression of 4 photosynthesis-related genes. These results could help clarify the molecular mechanisms underlying D. salina responses to the Yuncheng Salt Lake environment and lay a foundation for further utilization of this algal resource.
Key words:    Dunaliella salina    transcriptome analysis    de novo assembly    salt stress   
Received: 2020-04-18   Revised: 2020-06-08
PDF (4084 KB) Free
Print this page
Add to favorites
Email this article to others
Articles by Fan GAO
Articles by Fangru NAN
Articles by Jia FENG
Articles by Junping LÜ
Articles by Qi LIU
Articles by Xudong LIU
Articles by Shulian XIE
Afify A E M M R, Shalaby E A, Shanab S M M. 2010. Enhancement of biodiesel production from different species of algae. Grasas y Aceites, 61(4):416-422,
Ahmed R A, He M L, Aftab R A, Zheng S Y, Nagi M, Bakri R, Wang C H. 2017. Bioenergy application of Dunaliella salina SA 134 grown at various salinity levels for lipid production. Scientific Reports, 7:8118,
Alkayal F, Albion R L, Tillett R L, Hathwaik L T, Lemos M S, Cushman J C. 2010. Expressed sequence tag (EST) profiling in hyper saline shocked Dunaliella salina reveals high expression of protein synthetic apparatus components. Plant Science, 179(5):437-449,
Azachi M, Sadka A, Fisher M, Goldshlag P, Gokhman I, Zamir A. 2002. Salt induction of fatty acid elongase and membrane lipid modifications in the extreme halotolerant alga Dunaliella salina. Plant Physiology, 129(3):1320-1329,
BenMoussa-Dahmen I, Chtourou H, Rezgui F, Sayadi S, Dhouib A. 2016. Salinity stress increases lipid, secondary metabolites and enzyme activity in Amphora subtropica and Dunaliella sp. for biodiesel production. Bioresource Technology, 218:819-825,
Biasini M, Bienert S, Waterhouse A, Arnold K, Studer G, Schmidt T, Kiefer F, Cassarino T G, Bertoni M, Bordoli L, Schwede T. 2014. SWISS-MODEL:modelling protein tertiary and quaternary structure using evolutionary information. Nucleic Acids Research, 42(W1):W252-W258,
Chen X J, Wu M J, Jiang Y, Yang Y, Yan Y B. 2015. Dunaliella salina Hsp90 is halotolerant. International Journal of Biological Macromolecules, 75:418-425,
Chen Y X, Chen Y S, Shi C M, Huang Z B, Zhang Y, Li S K, Li Y, Ye J, Yu C, Li Z, Zhang X Q, Wang J, Yang H M, Fang L, Chen Q. 2018. SOAPnuke:a mapreduce acceleration-supported software for integrated quality control and preprocessing of high-throughput sequencing data. Gigascience, 7(1):gix120,
Cheng S F, Xian W F, Fu Y, Marin B, Keller J, Wu T, Sun W J, Li X L, Xu Y, Zhang Y, Wittek S, Reder T, Günther G, Gontcharov A, Wang S B, Li L Z, Liu X, Wang J, Yang H M, Xu X, Delaux P M, Melkonian B, Wong G K S, Melkonian M. 2019. Genomes of subaerial Zygnematophyceae provide insights into land plant evolution. Cell, 179(5):1057-1067.e14,
Davidi L, Katz A, Pick U. 2012. Characterization of major lipid droplet proteins from Dunaliella. Planta, 236:19-33,
Djebali S, Wucher V, Foissac S, Hitte C, Corre E, Derrien T. 2017. Bioinformatics pipeline for transcriptome sequencing analysis. Methods in Molecular Biology, 1468:201-219,
Emmerstorfer-Augustin A, Moser S, Pichler H. 2016. Screening for improved isoprenoid biosynthesis in microorganisms. Journal of Biotechnology, 235:112-120,
Fazeli M R, Tofighi H, Samadi N, Jamalifar H. 2006. Effects of salinity on β-carotene production by Dunaliella tertiolecta DCCBC26 isolated from the Urmia salt lake, north of Iran. Bioresource Technology, 97(18):2453-2456,
Fu X P, Wang D C, Yin X L, Du P C, Kan B. 2014. Time course transcriptome changes in Shewanella algae in response to salt stress. PLoS One, 9(5):e96001,
Gasteiger E, Gattiker A, Hoogland C, Ivanyi I, Appel R D, Bairoch A. 2003. ExPASy:the proteomics server for indepth protein knowledge and analysis. Nucleic Acids Research, 31(13):3784-3788,
Gierahn T M, Wadsworth Ⅱ M H, Hughes T K, Bryson B D, Butler A, Satija R, Fortune S, Love J C, Shalek A K. 2017. Seq-well:portable, low-cost RNA sequencing of single cells at high throughput. Nature Methods, 14(4):395-398,
Gong F H, Wang J, Li J. 2017. Isolation and characterization of peroxiredoxin 1 gene of Dunaliella salina. Gene, 635:39-45,
Götz S, García-Gómez J M, Terol J, Williams T D, Nagaraj S H, Nueda M J, Robles M, Talón M, Dopazo J, Conesa A. 2008. High-throughput functional annotation and data mining with the Blast2GO suite. Nucleic Acids Research, 36(10):3420-3435,
Han C, Li Q, Li X F, Zhang Z P, Huang J R. 2018. De novo assembly, characterization and annotation for the transcriptome of Sphaeroma terebrans and microsatellite marker discovery. Genes & Genomics, 40(2):167-176,
He Q H, Lin Y Q, Tan H, Zhou Y, Wen Y L, Gan J J, Li R W, Zhang Q L. 2020. Transcriptomic profiles of Dunaliella salina in response to hypersaline stress. BMC Genomics, 21(1):115,
He Z H, Qin K J, Wang Y, Zhao W. 1993. Biological resources in inland saline waters from Southern Shanxi, China Part I. Lake Xiao Chi. Journal of Dalian Fisheries College, 8(4):1-16, (in Chinese with English abstract)
Hong L, Liu J L, Midoun S Z, Miller P C. 2017. Transcriptome sequencing and annotation of the halophytic microalga Dunaliella salina. Journal of Zhejiang University-Science B, 18(10):833-844,
Hu L, Zheng T C, Cai M, Pan H T, Wang J, Zhang Q X. 2019. Transcriptome analysis during floral organ development provides insights into stamen petaloidy in Lagerstroemia speciosa. Plant Physiology and Biochemistry, 142:510-518,
Huang W C, Kim J D. 2017. Simultaneous cell disruption and lipid extraction in a microalgal biomass using a nonpolar tertiary amine. Bioresource Technology, 232:142-145,
Im S, Lee H N, Jung H S, Yang S, Park E J, Hwang M S, Jeong W J, Choi D W. 2017. Transcriptome-based identification of the desiccation response genes in marine red algae Pyropia tenera (Rhodophyta) and enhancement of abiotic stress tolerance by PtDRG2 in Chlamydomonas. Marine Biotechnology, 19(3):232-245,
Jin J P, Tian F, Yang D C, Meng Y Q, Kong L, Luo J C, Gao G. 2017. PlantTFDB 4.0:toward a central hub for transcription factors and regulatory interactions in plants. Nucleic Acids Research, 45(D1):D1040-1045,
Jin Y, Qian H, Yu M J. 2015. Phylogenetic structure of tree species across different life stages from seedlings to canopy trees in a subtropical evergreen broad-leaved forest. PLoS One, 10(6):e0131162,
Kawase Y, Imamura S, Tanaka K. 2017. A MYB-type transcription factor, MYB2, represses light-harvesting protein genes in Cyanidioschyzon merolae. FEBS Letters, 591(16):2439-2448,
Kumar L, Futschik M E. 2007. Mfuzz:a software package for soft clustering of microarray data. Bioinformation, 2(1):5-7,
Langdon W B. 2015. Performance of genetic programming optimised Bowtie2 on genome comparison and analytic testing (GCAT) benchmarks. Biodata Mining, 8:1,
Lee J, Heath L S, Grene R, Li S. 2019. Comparing time series transcriptome data between plants using a network module finding algorithm. Plant Methods, 15:61,
Li J, Lu Y M, Xue L X, Xie H. 2010. A structurally novel saltregulated promoter of duplicated carbonic anhydrase gene 1 from Dunaliella salina. Molecular Biology Reports, 37(2):1143-1154,
Li X Y, Yuan Y Z, Cheng D J, Gao J, Kong L Z, Zhao Q Y, Wei W, Sun Y H. 2018. Exploring stress tolerance mechanism of evolved freshwater strain Chlorella sp. S30 under 30 g/L salt. Bioresource Technology, 250:495-504,
Liang M H, Lu Y, Chen H H, Jiang J G. 2017. The saltregulated element in the promoter of lycopene β-cyclase gene confers a salt regulatory pattern in carotenogenesis of Dunaliella bardawil. Environmental Microbiology, 19(3):982-989,
Liang X, Qiao D R, Huang M, Yi X L, Bai L H, Xu H, Wei L, Zeng J, Cao Y. 2008. Identification of a gene encoding the light-harvesting chlorophyll a/b proteins of photosystem I in green alga Dunaliella salina. DNA Sequence, 19(2):137-145,
Mak S S T, Gopalakrishnan S, Carøe C, Geng C Y, Liu S L, Sinding M H S, Kuderna L F K, Zhang W W, Fu S J, Vieira F G, Germonpré M, Bocherens H, Fedorov S, Petersen B, Sicheritz-Pontén T, Marques-Bonet T, Zhang G J, Jiang H, Gilbert M T P. 2017. Comparative performance of the BGISEQ-500 vs Illumina HiSeq2500
sequencing platforms for palaeogenomic sequencing. Gigascience, 6(8):gix049,
Mansfeldt C B, Richter L V, Ahner B A, Cochlan W P, Richardson R E. 2016. Use of De novo transcriptome libraries to characterize a novel oleaginous marine Chlorella species during the accumulation of triacylglycerols. PLoS One, 11(2):e0147527,
Mao X M, Zhang Y, Wang X F, Liu J. 2020. Novel insights into salinity-induced lipogenesis and carotenogenesis in the oleaginous astaxanthin-producing alga Chromochloris zofingiensis:a multi-omics study. Biotechnology for Biofuels, 13:73,
Maza E. 2016. In papyro comparison of TMM (edgeR), RLE (DESeq2), and MRN normalization methods for a simple two-conditions-without-replicates RNA-Seq experimental design. Frontiers in Genetics, 7:164,
Miura R, Araki A, Minatoya M, Miyake K, Chen M L, Kobayashi S, Miyashita C, Yamamoto J, Matsumura T, Ishizuka M, Kubota T, Kishi R. 2019. An epigenomewide analysis of cord blood DNA methylation reveals sex-specific effect of exposure to bisphenol A. Scientific Reports, 9:12369,
Moriya Y, Itoh M, Okuda S, Yoshizawa A C, Kanehisa M. 2007. KAAS:an automatic genome annotation and pathway reconstruction server. Nucleic Acids Research, 35(S2):W182-W185,
Panahi B, Frahadian M, Dums J T, Hejazi M A. 2019. Integration of cross species RNA-Seq meta-analysis and machine-learning models identifies the most important salt stress-responsive pathways in microalga Dunaliella. Frontiers in Genetics, 10:752,
Pertea G, Huang X Q, Liang F, Antonescu V, Sultana R, Karamycheva S, Lee Y, White J, Cheung F, Parvizi B, Tsai J, Quackenbush J. 2003. TIGR gene indices clustering tools (TGICL):a software system for fast clustering of large EST datasets. Bioinformatics, 19(5):651-652,
Polle J E W, Barry K, Cushman J, Schmutz J, Tran D, Hathwaik L T, Yim W C, Jenkins J, McKie-Krisberg Z, Prochnik S, Lindquist E, Dockter R B, Adam C, Molina H, Bunkenborg J, Jin E, Buchheim M, Magnuson J. 2017. Draft nuclear genome sequence of the halophilic and beta-caroteneaccumulating green alga Dunaliella salina strain CCAP19/18. Genome Announcements, 5(43):e01105-17,
Puente-Sánchez F, Olsson S, Aguilera A. 2016. Comparative transcriptomic analysis of the response of Dunaliella acidophila (Chlorophyta) to short-term cadmium and chronic natural metal-rich water exposures. Microbial Ecology, 72(3):595-607,
Qv X Y, Jiang J G. 2013. Toxicity evaluation of two typical surfactants to Dunaliella bardawil, an environmentally tolerant alga. Environmental Toxicology and Chemistry, 32(2):426-433,
Ramos A A, Polle J, Tran D, Cushman J C, Jin E, Varela J C. 2011. The unicellular green alga Dunaliella salina Teod. as a model for abiotic stress tolerance:genetic advances and future perspectives. Algae, 26(1):3-20,
Rismani-Yazdi H, Haznedaroglu BZ, Bibby K, Peccia J. 2011. Transcriptome sequencing and annotation of the microalgae Dunaliella tertiolecta:pathway description and gene discovery for production of next-generation biofuels. BMC Genomics, 12:148,
Rodríguez-García A, Sola-Landa A, Barreiro C. 2017. RNASeq-Based comparative transcriptomics:RNA rreparation and bioinformatics. Methods in Molecular Biology, 1645:59-72,
Ruijter J M, Villalba A R, Hellemans J, Untergasser A, Van Den Hoff MJB. 2015. Removal of between-run variation in a multi-plate qPCR experiment. Biomolecular detection and quantification, 5:10-14,
Seddigh S, Darabi M. 2018. Functional, structural, and phylogenetic analysis of mitochondrial cytochrome b(cytb) in insects. Mitochondrial DNA Part A, 29(2):236-249,
Seppey M, Manni M, Zdobnov E M. 2019. BUSCO:assessing genome assembly and annotation completeness. Methods in Molecular Biology, 1962:227-245,
Shetty P, Gitau M M, Maróti G. 2019. Salinity stress responses and adaptation mechanisms in eukaryotic green microalgae. Cells, 8(12):1657,
Smith D R, Lee R W, Cushman J C, Magnuson J K, Tran D, Polle J E W. 2010. The Dunaliella salina organelle genomes:large sequences, inflated with intronic and intergenic DNA. BMC Plant Biology, 10:83,
Solovchenko A E, Selivanova E A, Chekanov K A, Sidorov R A, Nemtseva N V, Lobakova E S. 2015. Induction of secondary carotenogenesis in new halophile microalgae from the genus Dunaliella (Chlorophyceae). Biochemistry(Moscow), 80(11):1508-1513,
Song J Y, Yao H, Li Y, Li X W, Lin Y L, Liu C, Han J P, Xie C X, Chen S L. 2009. Authentication of the family polygonaceae in Chinese pharmacopoeia by DNA barcoding technique. Journal of Ethnopharmacology, 124(3):434-439,
Talebi A F, Tabatabaei M, Mohtashami S K, Tohidfar M, Moradi F. 2013. Comparative salt stress study on intracellular ion concentration in marine and salt-adapted freshwater strains of microalgae. Notulae Scientia Biologicae, 5(3):309-315,
Tian Y Y, Bai S, Dang Z H, Hao J F, Zhang J, Hasi A. 2019. Genome-wide identification and characterization of long non-coding RNAs involved in fruit ripening and the climacteric in Cucumis melo. BMC Plant Biology, 19:369,
Van De Poel B, Cooper E D, Van Der Straeten D, Chang C, Delwiche C F. 2016. Transcriptome profiling of the green alga Spirogyra pratensis (Charophyta) suggests an ancestral role for ethylene in cell wall metabolism, photosynthesis, and abiotic stress responses. Plant Physiology, 172(1):533-545,
Vasilikiotis C, Melis, A. 1994. Photosystem Ⅱ reaction center damage and repair cycle:chloroplast acclimation strategy to irradiance stress. Proceedings of the National Academy of Sciences of the United States of America, 91(15):7222-7226,
Wang F P, Feng J, Wang J, Li B, Xie S L. 2014. Phylogenetic and morphological investigation of a Dunaliella strain isolated from Yuncheng Salt Lake, China. Acta Geologica Sinica (English Edition), 88(Supp. l):106-107,
Wang M X, Zhang Y X, Guo P Y. 2017. Effect of florfenicol and thiamphenicol exposure on the photosynthesis and antioxidant system of Microcystis flos-aquae. Aquatic Toxicology, 186:67-76,
Wang P, Wang Z N, Dou Y C, Zhang X X, Wang M Y, Tian X M. 2013. Genome-wide identification and analysis of membrane-bound O-acyltransferase (MBOAT) gene family in plants. Planta, 238(5):907-922,
Wang Y, Cong Y T, Wang Y H, Guo Z H, Yue J R, Xing Z Y, Gao X N, Chai X J. 2019. Identification of early salinity stress-responsive proteins in Dunaliella salina by isobaric tags for relative and absolute quantitation (iTRAQ)-based quantitative proteomic analysis. International Journal of Molecular Sciences, 20(3):599,
Wei S J, Bian Y Y, Zhao Q, Chen S X, Mao J W, Song C X, Cheng K, Xiao Z, Zhang C F, Ma W M, Zou H F, Ye M L, Dai S J. 2017. Salinity-induced palmella formation mechanism in halotolerant algae Dunaliella salina revealed by quantitative proteomics and phosphoproteomics. Frontiers in Plant Science, 8:810,
Wheeler D L, Church D M, Edgar R, Federhen S, Helmberg W, Madden T L, Pontius J U, Schuler G D, Schriml L M, Sequeira E, Suzek T O, Tatusova T A, Wagner L. 2004. Database resources of the National Center for Biotechnology Information:update. Nucleic Acids Research, 32(S1):D35-D40,
Wichuk K, Brynjólfsson S, Fu W Q. 2014. Biotechnological production of value-added carotenoids from microalgae:emerging technology and prospects. Bioengineered, 5(3):204-208,
Wu Q, Lan Y H, Cao X Y, Yao H Y, Qiao D R, Xu H, Cao Y. 2019. Characterization and diverse evolution patterns of glycerol-3-phosphate dehydrogenase family genes in Dunaliella salina. Gene, 710:161-169,
Wu X L, Xiong E H, Wang W, Scali M, Cresti M. 2014. Universal sample preparation method integrating trichloroacetic acid/acetone precipitation with phenol extraction for crop proteomic analysis. Nature Protocols, 9(2):362-374,
Xie S L, Li Y H, Li Z. 1998. The species composition and distribution of Euglenophyta in salt lakes area in Yuncheng. Acta Hydrobiologica Sinica, 22(1):33-38. (in Chinese with English abstract)
Yao L N, Tan K W M, Tan T W, Lee Y K. 2017. Exploring the transcriptome of non-model oleaginous microalga Dunaliella tertiolecta through high-throughput sequencing and high performance computing. BMC Bioinformatics, 18:122,
Yao L N, Tan T W, Ng Y K, Ban K H K, Shen H, Lin H X, Lee Y K. 2015. RNA-Seq transcriptomic analysis with Bag2D software identifies key pathways enhancing lipid yield in a high lipid-producing mutant of the non-model green alga Dunaliella tertiolecta. Biotechnology for Biofuels, 8:191,
Yuan P P, Yin S, Han D, Zhang W J, Cui H L. 2015. Halorientalis brevis sp. nov., isolated from an inland salt lake of China. Current Microbiology, 71(3):382-386,
Zhang S, Li X R, Xu H, Cao Y, Ma S H, Cao Y, Qiao D R. 2014. Molecular cloning and functional characterization of MnSOD from Dunaliella salina. Journal of Basic Microbiology, 54(5):438-447,
Zhou L Q, Liu Z H, Dong Y H, Sun X J, Wu B, Yu T, Zheng Y X, Yang A G, Zhao Q, Zhao D. 2019. Transcriptomics analysis revealing candidate genes and networks for sex differentiation of yesso scallop (Patinopecten yessoensis). BMC Genomics, 20:671,
Zhou Y. 2013. The Expression Analylsis of Genes Encoding Light-Harvesting Chlorophyll a/b-Binding Proteins in Zostera marina L. in Differernt Environment. Ocean University of China, Qingdao.

Related Articles:
1.CONG Yuting, WANG Yuan, YUE Jinrong, XING Zhenyu, GAO Xiangnan, CHAI Xiaojie.Expression, purification, and subcellular localization of phospholipase C in Dunaliella salina[J]. Journal of Oceanology and Limnology, 2019,37(4): 1363-1371
Copyright © Haiyang Xuebao