Cre-mediated marker gene removal for production of biosafe commercial oilseed rape

• Autorzy: Boszoradova E. , Matuskova I. , Libantova J. , Zimova M. , Moravcikova J.
• Języki publikacji: EN

Abstrakty

EN

The Cre/loxP-based self-excision represents a promising strategy for removal of hazardous transgene(s) from genomes of targeted crops prior to their introduction into environment. Here, we applied the Cre/loxP self-excision strategy in which the cre recombinase gene is driven by the embryo-specific CRUC promoter from Arabidopsis thaliana. Besides, the Cre/loxP cassette, the T-DNA also consisted of the β-glucuronidase gene, controlled by the double dCaMV 35S promoter and the selectable neomycin phosphotransferase gene; the latter was aimed to be removed from the transgenic genome. The Cre/loxP self-excision cassette was introduced into genomes of four commercial oilseed rape cultivars Haydn, Heros, Hunter and Topas (Brassica napus L.) via Agrobacterium tumefaciens. Transgenic T₀ plants were regenerated from all cultivars. Their detailed molecular analyses revealed premature activation of the CRUC promoter that resulted in partial excision of the nptII gene. Progenies of four self-pollinated T₀ lines were further analysed, and the data confirmed complete excision event in 5 out of 105 (4.8%) of T₁ transgenic oilseed rape plants. The excision efficiency does not seem to depend on the target cultivar. However, the poor transformation efficiency of rapeseed and the limited specificity of the CRUC promoter are clearly the bottleneck of this approach, and the feasibility of (tissue-specific) self-excision of selectable marker gene from genomes of each commercial rapeseed variety adds to their perspective to cope the increasing negative impacts of climate changes.

• Wydawca: -
• Czasopismo: Acta Physiologiae Plantarum
• Rocznik: 2019
• Tom: 41
• Numer: 06
• Opis fizyczny: Article 73 [8p.], fig.,ref.

Twórcy

autor

Boszoradova E.

• Plant Science and Biodiversity Center, Institute of Plant Genetics and Biotechnology, Slovak Academy of Sciences, Akademicka 2, P.O. Box 39A, 950 07 Nitra, Slovak Republic

autor

Matuskova I.

• Department of Ecochemistry and Radioecology, University of Cyril and Methodius, J. Herdu 2, 917 01 Trnava, Slovak Republic

autor

Libantova J.

• Plant Science and Biodiversity Center, Institute of Plant Genetics and Biotechnology, Slovak Academy of Sciences, Akademicka 2, P.O. Box 39A, 950 07 Nitra, Slovak Republic

autor

Zimova M.

• Department of Botany and Genetics, Faculty of Natural Sciences, Constantine Philosopher University, Nábrezie mladeze 91, 94974 Nitra, Slovak Republic

autor

Moravcikova J.

• Department of Biotechnology, University of Cyril and Methodius, J. Herdu 2, 917 01 Trnava, Slovak Republic

Bibliografia

• Bai XQ, Wang QY, Chu CC (2008) Excision of a selective marker in transgenic rice using a novel Cre/loxP system controlled by a floral specific promoter. Transgenic Res 17:1035–1043
• Bailleul D, Ollier S, Lecomte J (2016) Genetic diversity of oilseed rape fields and feral populations in the context of coexistence with GM crops. PLoS One 11:e0158403
• Becerra C, Puigdomenech P, Vicient CM (2006) Computational and experimental analysis identifies Arabidopsis genes specifically expressed during early seed development. BMC Genom 7:38
• Bondarenko VA, Jiang YI, Studitsky VM (2003) Rationally designed insulator-like elements can block enhancer action in vitro. EMBO J 22:4728–4737
• Boszoradova E, Libantova J, Matusikova I, Poloniova Z, Jopcik M, Berenyi M, Moravcikova J (2011) Agrobacterium-mediated genetic transformation of economically important oilseed rape cultivars. Plant Cell Tissue Organ Cult 107:317–323
• Boszoradova E, Libantova J, Matsuikova I, Moravcikova J (2014) Application of Arabidopsis tissue-specific CRUC promoter in the Cre/loxP self-excision strategy for generation of marker-free oilseed rape: potential advantages and drawbacks. Acta Physiol Plant 36:1399–1409
• Chakraborti D, Sarkar A, Mondal HA, Schuermann D, Hohn B, Sarmah BK, Das S (2008) Cre/lox system to develop selectable marker free transgenic tobacco plants conferring resistance against sap sucking homopteran insect. Plant Cell Rep 27:1623–1633
• Chen J, Greenblatt IM, Dellaporta SL (1992) Molecular analysis of ac transposition and DNA-replication. Genetics 130:665–676
• Chen H, Luo J, Zheng P, Zhang XB, Zhang CC, Li XY, Wang MG, Huang YQ, Liu XJ, Jan M, Liu YJ, Hu PS, Tu JM (2017) Application of Cre-lox gene switch to limit the Cry expression in rice green tissues. Sci Rep 7:14505
• Chong-Perez B, Reyes M, Rojas L, Ocana B, Ramos A, Kosky RG, Angenon G (2013) Excision of a selectable marker gene in transgenic banana using a Cre/lox system controlled by an embryo specific promoter. Plant Mol Biol 83:143–152
• Éva C, Teglas F, Zelenyanszki H, Tamas C, Juhasz A, Meszaros K, Tamas L (2018) Cold inducible promoter driven Cre-lox system proved to be highly efficient for marker gene excision in transgenic barley. J Biotechnol 265:15–24
• Gidoni D, Srivastava V, Carmi N (2008) Site-specific excisional recombination strategies for elimination of undesirable transgenes from crop plants. In Vitro Cell Dev Biol Plant 44:457–467
• Gilbertson L (2003) Cre-lox recombination: Cre-ative tools for plant biotechnology. Trends Biotechnol 21:550–555
• Goldsbrough AP, Lastrella CN, Yoder JI (1993) Transposition mediated repositioning and subsequent elimination of marker genes from transgenic tomato. Bio-Technology 11:1286–1292
• Hamzeh S, Motallebi M, Zamani MR (2016) Efficient seed-specifically regulated autoexcision of marker gene (nptII) with inducible expression of interest gene in transgenic Nicotiana tabacum. Turk J Biol 40:1–11
• Hoenicka H, Lehnhardt D, Nunna S, Reinhardt R, Jeltsch A, Briones V, Fladung M (2016) Level of tissue differentiation influences the activation of a heat-inducible flower-specific system for genetic containment in poplar (Populus tremula L.). Plant Cell Rep 35:369–384
• Hoglund AS, Rodin J, Larsson E, Rask L (1992) Distribution of napin and cruciferin in developing rape seed embryos. Plant Physiol 98:509–515
• ISAAA (2018) International service for the acquisition of agri-biotech applications GM approval database. http://www.isaaa.org/gmapprovaldatabase/default.asp. Accessed 15 June 2018
• Jefferson RA, Kavanagh TA, Bevan MW (1987) Gus fusions—beta-glucuronidase as a sensitive and versatile gene fusion marker in higher-plants. EMBO J 6:3901–3907
• Kirik A, Salomon S, Puchta H (2000) Species-specific double-strand break repair and genome evolution in plants. EMBO J 19:5562–5566
• Kopertekh L, Broer I, Schiemann J (2009) Developmentally regulated site-specific marker gene excision in transgenic B. napus plants. Plant Cell Rep 28:1075–1083
• Kopertekh L, Schulze K, Frolov A, Strack D, Broer I, Schiemann J (2010) Cre-mediated seed-specific transgene excision in tobacco. Plant Mol Biol 72:597–605
• Kopertekh L, Krebs E, Guzmann F (2018) Improvement of conditional Cre-lox system through application of the regulatory sequences from Cowpea mosaic virus. Plant Biotechnol Rep 12:127–137
• Li ZS, Xing AQ, Moon BP, Burgoyne SA, Guida AD, Liang HL, Lee C, Caster CS, Barton JE, Klein TM, Falco SC (2007) A Cre/loxP-mediated self-activating gene excision system to produce marker gene free transgenic soybean plants. Plant Mol Biol 65:329–341
• Luo KM, Duan H, Zhao DG, Zheng XL, Deng W, Chen YQ, Stewart CN, McAvoy R, Jiang XN, Wu YH, He AG, Pei Y, Li Y (2007) ‘GM-gene-deletor’: fused loxP-FRT recognition sequences dramatically improve the efficiency of FLP or CRE recombinase on transgene excision from pollen and seed of tobacco plants. Plant Biotechnol J 5:263–274
• Ma BG, Duan XY, Niu JX, Ma C, Hao QN, Zhang LX, Zhang HP (2009) Expression of stilbene synthase gene in transgenic tomato using salicylic acid-inducible Cre/loxP recombination system with self-excision of selectable marker. Biotechnol Lett 31:163–169
• Marjanac G, De Paepe A, Peck I, Jacobs A, De Buck S, Depicker A (2008) Evaluation of CRE-mediated excision approaches in Arabidopsis thaliana. Transgenic Res 17:239–250
• Meszaros K, Eva C, Kiss T, Banyai J, Kiss E, Teglas F, Lang L, Karsai I, Tamas L (2015) Generating marker-free transgenic wheat using minimal gene cassette and cold-inducible Cre/Lox system. Plant Mol Biol Rep 33:1221–1231
• Mietkiewska E, Dees D, Nap JP (2000) Seed specific GUS gene expression directed by the Arabidopsis cruciferin C promoter in tobacco and Arabidopsis. In: Session plant lipid metabolisms, 6th international congress of plant molecular biology, ISPMB, June 18–24, Quebec
• Mlynarova L, Conner AJ, Nap JP (2006) Directed microspore-specific recombination of transgenic alleles to prevent pollen-mediated transmission of transgenes. Plant Biotechnol J 4:445–452
• Moravcikova J, Vaculkova E, Bauer M, Libantova J (2008) Feasibility of the seed specific cruciferin C promoter in the self excision Cre/loxP strategy focused on generation of marker-free transgenic plants. Theor Appl Genet 117:1325–1334
• Murashige I, Skoog F (1962) A revised medium for rapid growth and bioassays with tobacco tissue cultures. Plant Physiol 15:473–497
• Pfaffl MW (2001) A new mathematical model for relative quantification in real-time RT-PCR. Nucleic Acids Res 29:2002–2007
• Poloniova Z, Jopcik M, Matusikova I, Libantova J, Moravcikova J (2015) The pollen- and embryo-specific Arabidopsis DLL promoter bears good potential for application in marker-free Cre/loxP self-excision strategy. Plant Cell Rep 34:469–481
• Pradhan S, Chakraborty A, Sikdar N, Chakraborty S, Bhattacharyya J, Mitra J, Manna A, Gupta SD, Sen SK (2016) Marker-free transgenic rice expressing the vegetative insecticidal protein (Vip) of Bacillus thuringiensis shows broad insecticidal properties. Planta 244:789–804
• Puchta H (2005) The repair of double-strand breaks in plants: mechanisms and consequences for genome evolution. J Exp Bot 56:1–14
• Qiu CX, Sangha JS, Song FS, Zhou ZY, Yin A, Gu KY, Tian DS, Yang JB, Yin ZC (2010) Production of marker-free transgenic rice expressing tissue-specific Bt gene. Plant Cell Rep 29:1097–1107
• Roy S (2014) Maintenance of genome stability in plants: repairing DNA double strand breaks and chromatin structure stability. Front Plant Sci 5:487
• Science for Environment Policy (2017) European Commission DG Environment News Alert Service, edited by SCU. The University of the West of England, Bristol. http://ec.europa.eu/environment/integration/research/newsalert/pdf/oilseed_rape_genes_transfer_inside_to_outside_crop_fields_gm_risk_assessment_485na1_en.pdf. Accessed 15 June 2018
• Sengupta S, Chakraborti D, Mondal HA, Das S (2010) Selectable antibiotic resistance marker gene-free transgenic rice harbouring the garlic leaf lectin gene exhibits resistance to sap-sucking planthoppers. Plant Cell Rep 29:261–271
• Siebert R, Puchta H (2002) Efficient repair of genomic double-strand breaks by homologous recombination between directly repeated sequences in the plant genome. Plant Cell 14:1121–1131
• Thelen JJ, Ohlrogge JB (2002) Metabolic engineering of fatty acid biosynthesis in plants. Metab Eng 4:12–21
• Thompson JD, Higgins DG, Gibson TJ (1994) CLUSTAL-W—improving the sensitivity of progressive multiple sequence alignment through sequence weighting, position-specific gap penalties and weight matrix choice. Nucleic Acids Res 22:4673–4680
• Vaculkova E, Moravcikova J, Matusikova I, Bauer M, Libantova J (2007) A modified low copy number binary vector pUN for Agrobacterium-mediated plant transformation. Biol Plant 51:538–540
• Vigeolas H, Waldeck P, Zank T, Geigenberger P (2007) Increasing seed oil content in oil-seed rape (Brassica napus L.) by over-expression of a yeast glycerol-3-phosphate dehydrogenase under the control of a seed-specific promoter. Plant Biotechnol J 5:431–441
• Zhang Y, Liu H, Li B, Zhang JT, Li YZ, Zhang HX (2009) Generation of selectable marker-free transgenic tomato resistant to drought, cold and oxidative stress using the Cre/loxP DNA excision system. Transgenic Res 18:607–619
• Zheng Y, Pan Y, Li JH, Zhou Y, Pan YL, Ding YM, Su CG, Zhang XG (2016) Visible marker excision via heat-inducible Cre/LoxP system and Ipt selection in tobacco. In Vitro Cell Dev Biol Plant 52:492–499

Identyfikatory

DOI

10.1007/s11738-019-2865-2