Intron-containing algal transgenes mediate efficient recombinant gene expression in the green microalga Chlamydomonas reinhardtii

Baier T, Wichmann J, Kruse O, Lauersen KJ (2018)
Nucleic Acids Research 46(13): 6909-6919.

Download
OA 1.84 MB
Zeitschriftenaufsatz | Veröffentlicht | Englisch
Volltext vorhanden für diesen Nachweis
Abstract / Bemerkung
Among green freshwater microalgae, Chlamydomonas reinhardtii has the most comprehensive and developed molecular toolkit, however, advanced genetic and metabolic engineering driven from the nuclear genome is generally hindered by inherently low transgene expression levels. Progressive strain development and synthetic promoters have improved the capacity of transgene expression; however, the responsible regulatory mechanisms are still not fully understood. Here, we elucidate the sequence specific dynamics of native regulatory element insertion into nuclear transgenes. Systematic insertions of the first intron of the ribulose-1,5-bisphosphate carboxylase/oxygenase small subunit 2 (rbcS2i1) throughout codon-optimized coding sequences (CDS) generates optimized algal transgenes which express reliably in C. reinhardtii. The optimal rbcS2i1 insertion site for efficient splicing was systematically determined and improved gene expression rates were shown using a codon-optimized sesquiterpene synthase CDS. Sequential insertions of rbcS2i1 were found to have a step-wise additive effect on all levels of transgene expression, which is likely correlated to a synergy of transcriptional machinery recruitment and mimicking the short average exon lengths natively found in the C. reinhardtii genome. We further demonstrate the value of this optimization with five representative transgene examples and provide guidelines for the design of any desired sequence with this strategy.
Erscheinungsjahr
Zeitschriftentitel
Nucleic Acids Research
Band
46
Ausgabe
13
Seite(n)
6909-6919
ISSN
eISSN
PUB-ID

Zitieren

Baier T, Wichmann J, Kruse O, Lauersen KJ. Intron-containing algal transgenes mediate efficient recombinant gene expression in the green microalga Chlamydomonas reinhardtii. Nucleic Acids Research. 2018;46(13):6909-6919.
Baier, T., Wichmann, J., Kruse, O., & Lauersen, K. J. (2018). Intron-containing algal transgenes mediate efficient recombinant gene expression in the green microalga Chlamydomonas reinhardtii. Nucleic Acids Research, 46(13), 6909-6919. doi:10.1093/nar/gky532
Baier, T., Wichmann, J., Kruse, O., and Lauersen, K. J. (2018). Intron-containing algal transgenes mediate efficient recombinant gene expression in the green microalga Chlamydomonas reinhardtii. Nucleic Acids Research 46, 6909-6919.
Baier, T., et al., 2018. Intron-containing algal transgenes mediate efficient recombinant gene expression in the green microalga Chlamydomonas reinhardtii. Nucleic Acids Research, 46(13), p 6909-6919.
T. Baier, et al., “Intron-containing algal transgenes mediate efficient recombinant gene expression in the green microalga Chlamydomonas reinhardtii”, Nucleic Acids Research, vol. 46, 2018, pp. 6909-6919.
Baier, T., Wichmann, J., Kruse, O., Lauersen, K.J.: Intron-containing algal transgenes mediate efficient recombinant gene expression in the green microalga Chlamydomonas reinhardtii. Nucleic Acids Research. 46, 6909-6919 (2018).
Baier, Thomas, Wichmann, Julian, Kruse, Olaf, and Lauersen, Kyle J. “Intron-containing algal transgenes mediate efficient recombinant gene expression in the green microalga Chlamydomonas reinhardtii”. Nucleic Acids Research 46.13 (2018): 6909-6919.
Alle Dateien verfügbar unter der/den folgenden Lizenz(en):
Creative Commons Namensnennung-Nicht kommerziell 4.0 International (CC BY-NC 4.0):
Volltext(e)
Access Level
OA Open Access
Zuletzt Hochgeladen
2018-07-31T10:23:46Z

2 Zitationen in Europe PMC

Daten bereitgestellt von Europe PubMed Central.

Selectable Markers and Reporter Genes for Engineering the Chloroplast of Chlamydomonas reinhardtii.
Esland L, Larrea-Alvarez M, Purton S., Biology (Basel) 7(4), 2018
PMID: 30309004
Prospects on the Use of Schizochytrium sp. to Develop Oral Vaccines.
Ramos-Vega A, Rosales-Mendoza S, Bañuelos-Hernández B, Angulo C., Front Microbiol 9(), 2018
PMID: 30410471

55 References

Daten bereitgestellt von Europe PubMed Central.

Genetic improvement of the microalga Phaeodactylum tricornutum for boosting neutral lipid accumulation.
Xue J, Niu YF, Huang T, Yang WD, Liu JS, Li HY., Metab. Eng. 27(), 2014
PMID: 25447640
Genetic tools and techniques for Chlamydomonas reinhardtii.
Mussgnug JH., Appl. Microbiol. Biotechnol. 99(13), 2015
PMID: 26025017
Towards developing algal synthetic biology.
Scaife MA, Smith AG., Biochem. Soc. Trans. 44(3), 2016
PMID: 27284033
Nuclear transformation and functional gene expression in the oleaginous microalga Monoraphidium neglectum.
Jaeger D, Hubner W, Huser T, Mussgnug JH, Kruse O., J. Biotechnol. 249(), 2017
PMID: 28302588
Chlamydomonas as a model organism
Harris E.H.., 2001
Algal sensory photoreceptors
Hegemann P., Fuhrmann M., Kateriya S.., 2001
Epigenetic silencing of a foreign gene in nuclear transformants of Chlamydomonas
Cerutti H.., 1997
Generation of Chlamydomonas strains that efficiently express nuclear transgenes.
Neupert J, Karcher D, Bock R., Plant J. 57(6), 2008
PMID: 19036032
Synthetic promoters capable of driving robust nuclear gene expression in the green alga Chlamydomonas reinhardtii
Scranton M.A., Ostrand J.T., Georgianna D.R., Lofgren S.M., Li D., Ellis R.C., Carruthers D.N., Dräger A., Masica D.L., Mayfield S.P.., 2016
Dissecting the contributions of GC content and codon usage to gene expression in the model alga Chlamydomonas reinhardtii.
Barahimipour R, Strenkert D, Neupert J, Schroda M, Merchant SS, Bock R., Plant J. 84(4), 2015
PMID: 26402748
Enhancing heterologous expression in Chlamydomonas reinhardtii by transcript sequence optimization.
Weiner I, Atar S, Schweitzer S, Eilenberg H, Feldman Y, Avitan M, Blau M, Danon A, Tuller T, Yacoby I., Plant J. 94(1), 2018
PMID: 29383789
Efficient foreign gene expression in Chlamydomonas reinhardtii mediated by an endogenous intron
Lumbreras V., Stevens D.R., Purton S.., 1998
Targeted expression of nuclear transgenes in Chlamydomonas reinhardtii with a versatile, modular vector toolkit.
Lauersen KJ, Kruse O, Mussgnug JH., Appl. Microbiol. Biotechnol. 99(8), 2015
PMID: 25586579
The Chlamydomonas genome reveals the evolution of key animal and plant functions.
Merchant SS, Prochnik SE, Vallon O, Harris EH, Karpowicz SJ, Witman GB, Terry A, Salamov A, Fritz-Laylin LK, Marechal-Drouard L, Marshall WF, Qu LH, Nelson DR, Sanderfoot AA, Spalding MH, Kapitonov VV, Ren Q, Ferris P, Lindquist E, Shapiro H, Lucas SM, Grimwood J, Schmutz J, Cardol P, Cerutti H, Chanfreau G, Chen CL, Cognat V, Croft MT, Dent R, Dutcher S, Fernandez E, Fukuzawa H, Gonzalez-Ballester D, Gonzalez-Halphen D, Hallmann A, Hanikenne M, Hippler M, Inwood W, Jabbari K, Kalanon M, Kuras R, Lefebvre PA, Lemaire SD, Lobanov AV, Lohr M, Manuell A, Meier I, Mets L, Mittag M, Mittelmeier T, Moroney JV, Moseley J, Napoli C, Nedelcu AM, Niyogi K, Novoselov SV, Paulsen IT, Pazour G, Purton S, Ral JP, Riano-Pachon DM, Riekhof W, Rymarquis L, Schroda M, Stern D, Umen J, Willows R, Wilson N, Zimmer SL, Allmer J, Balk J, Bisova K, Chen CJ, Elias M, Gendler K, Hauser C, Lamb MR, Ledford H, Long JC, Minagawa J, Page MD, Pan J, Pootakham W, Roje S, Rose A, Stahlberg E, Terauchi AM, Yang P, Ball S, Bowler C, Dieckmann CL, Gladyshev VN, Green P, Jorgensen R, Mayfield S, Mueller-Roeber B, Rajamani S, Sayre RT, Brokstein P, Dubchak I, Goodstein D, Hornick L, Huang YW, Jhaveri J, Luo Y, Martinez D, Ngau WC, Otillar B, Poliakov A, Porter A, Szajkowski L, Werner G, Zhou K, Grigoriev IV, Rokhsar DS, Grossman AR., Science 318(5848), 2007
PMID: 17932292
Comparative genomic analysis of retrogene repertoire in two green algae Volvox carteri and Chlamydomonas reinhardtii.
Jakalski M, Takeshita K, Deblieck M, Koyanagi KO, Makalowska I, Watanabe H, Makalowski W., Biol. Direct 11(), 2016
PMID: 27487948
Reconstruction of the lipid metabolism for the microalga Monoraphidium neglectum from its genome sequence reveals characteristics suitable for biofuel production.
Bogen C, Al-Dilaimi A, Albersmeier A, Wichmann J, Grundmann M, Rupp O, Lauersen KJ, Blifernez-Klassen O, Kalinowski J, Goesmann A, Mussgnug JH, Kruse O., BMC Genomics 14(), 2013
PMID: 24373495
Chromosome scale genome assembly and transcriptome profiling of Nannochloropsis gaditana in nitrogen depletion.
Corteggiani Carpinelli E, Telatin A, Vitulo N, Forcato C, D'Angelo M, Schiavon R, Vezzi A, Giacometti GM, Morosinotto T, Valle G., Mol Plant 7(2), 2013
PMID: 23966634
The Phaeodactylum genome reveals the evolutionary history of diatom genomes.
Bowler C, Allen AE, Badger JH, Grimwood J, Jabbari K, Kuo A, Maheswari U, Martens C, Maumus F, Otillar RP, Rayko E, Salamov A, Vandepoele K, Beszteri B, Gruber A, Heijde M, Katinka M, Mock T, Valentin K, Verret F, Berges JA, Brownlee C, Cadoret JP, Chiovitti A, Choi CJ, Coesel S, De Martino A, Detter JC, Durkin C, Falciatore A, Fournet J, Haruta M, Huysman MJ, Jenkins BD, Jiroutova K, Jorgensen RE, Joubert Y, Kaplan A, Kroger N, Kroth PG, La Roche J, Lindquist E, Lommer M, Martin-Jezequel V, Lopez PJ, Lucas S, Mangogna M, McGinnis K, Medlin LK, Montsant A, Oudot-Le Secq MP, Napoli C, Obornik M, Parker MS, Petit JL, Porcel BM, Poulsen N, Robison M, Rychlewski L, Rynearson TA, Schmutz J, Shapiro H, Siaut M, Stanley M, Sussman MR, Taylor AR, Vardi A, von Dassow P, Vyverman W, Willis A, Wyrwicz LS, Rokhsar DS, Weissenbach J, Armbrust EV, Green BR, Van de Peer Y, Grigoriev IV., Nature 456(7219), 2008
PMID: 18923393
A catalogue of splice junction sequences.
Mount SM., Nucleic Acids Res. 10(2), 1982
PMID: 7063411
Effect of intron A from human cytomegalovirus (Towne) immediate-early gene on heterologous expression in mammalian cells.
Chapman BS, Thayer RM, Vincent KA, Haigwood NL., Nucleic Acids Res. 19(14), 1991
PMID: 1650459
Strategies to facilitate transgene expression in Chlamydomonas reinhardtii.
Eichler-Stahlberg A, Weisheit W, Ruecker O, Heitzer M., Planta 229(4), 2009
PMID: 19127370
An optimized, chemically regulated gene expression system for Chlamydomonas.
Ferrante P, Catalanotti C, Bonente G, Giuliano G., PLoS ONE 3(9), 2008
PMID: 18787710
A gene-within-a-gene Cas9/sgRNA hybrid construct enables gene editing and gene replacement strategies in Chlamydomonas reinhardtii
Jiang W.Z., Weeks D.P.., 2017
Efficient phototrophic production of a high-value sesquiterpenoid from the eukaryotic microalga Chlamydomonas reinhardtii.
Lauersen KJ, Baier T, Wichmann J, Wordenweber R, Mussgnug JH, Hubner W, Huser T, Kruse O., Metab. Eng. 38(), 2016
PMID: 27474353
High-frequency nuclear transformation of Chlamydomonas reinhardtii.
Kindle KL., Proc. Natl. Acad. Sci. U.S.A. 87(3), 1990
PMID: 2105499
Single-step method of RNA isolation by acid guanidinium extraction
Chomczynski P.., 1987
Widespread distribution of a unique marine protistan lineage.
Cuvelier ML, Ortiz A, Kim E, Moehlig H, Richardson DE, Heidelberg JF, Archibald JM, Worden AZ., Environ. Microbiol. 10(6), 2008
PMID: 18341584
Form-determining function of the genes required for the assembly of the head of bacteriophage T4.
Laemmli UK, Molbert E, Showe M, Kellenberger E., J. Mol. Biol. 49(1), 1970
PMID: 5450520
Fast and sensitive colloidal Coomassie G-250 staining for proteins in polyacrylamide gels
Dyballa N., Metzger S.., 2009
RNAstructure: Web servers for RNA secondary structure prediction and analysis.
Bellaousov S, Reuter JS, Seetin MG, Mathews DH., Nucleic Acids Res. 41(Web Server issue), 2013
PMID: 23620284
Genome-wide analysis of alternative splicing in Chlamydomonas reinhardtii.
Labadorf A, Link A, Rogers MF, Thomas J, Reddy AS, Ben-Hur A., BMC Genomics 11(), 2010
PMID: 20163725

Harris E.H., Stern D., Witman G.., 2009
Robust expression of heterologous genes by selection marker fusion system in improved Chlamydomonas strains.
Kong F, Yamasaki T, Kurniasih SD, Hou L, Li X, Ivanova N, Okada S, Ohama T., J. Biosci. Bioeng. 120(3), 2015
PMID: 25660568
Escherichia coli K-12: a cooperatively developed annotation snapshot--2005.
Riley M, Abe T, Arnaud MB, Berlyn MK, Blattner FR, Chaudhuri RR, Glasner JD, Horiuchi T, Keseler IM, Kosuge T, Mori H, Perna NT, Plunkett G 3rd, Rudd KE, Serres MH, Thomas GH, Thomson NR, Wishart D, Wanner BL., Nucleic Acids Res. 34(1), 2006
PMID: 16397293
Engineering Yarrowia lipolytica for production of medium-chain fatty acids.
Rutter CD, Zhang S, Rao CV., Appl. Microbiol. Biotechnol. 99(17), 2015
PMID: 26129951
Sandalwood fragrance biosynthesis involves sesquiterpene synthases of both the terpene synthase (TPS)-a and TPS-b subfamilies, including santalene synthases.
Jones CG, Moniodis J, Zulak KG, Scaffidi A, Plummer JA, Ghisalberti EL, Barbour EL, Bohlmann J., J. Biol. Chem. 286(20), 2011
PMID: 21454632
The diverse sesquiterpene profile of patchouli, Pogostemon cablin, is correlated with a limited number of sesquiterpene synthases.
Deguerry F, Pastore L, Wu S, Clark A, Chappell J, Schalk M., Arch. Biochem. Biophys. 454(2), 2006
PMID: 16970904

Export

Markieren/ Markierung löschen
Markierte Publikationen

Open Data PUB

Web of Science

Dieser Datensatz im Web of Science®

Quellen

PMID: 30053227
PubMed | Europe PMC

Suchen in

Google Scholar