caa a22 4500 606208100 CHVBK 20210128101020.0 cr unu---uuuuu 210128e20151001xx s 000 0 eng 10.1007/s00425-015-2369-2 doi (NATIONALLICENCE)springer-10.1007/s00425-015-2369-2 Homologous sense and antisense expression of a gene in Dunaliella tertiolecta [Elektronische Daten] [Jing Zhang, Hancheol Jeon, Sang Sim, Yew Lee, EonSeon Jin] Main conclusion : Dunaliella transformation using antisense and sense technology developed in this study will provide powerful tools for functional analysis and pathway-specific metabolic engineering in Dunaliella for industrial applications. Our aim was to investigate the potential of antisense expression and overexpression of a specific gene, the carotenoid biosynthesis-related (CBR) gene, in the microalgae Dunaliella. DNA amplified from sense and antisense vector constructs was used to transform Dunaliella tertiolecta. The Gateway vector for plant transformation was used to make an expression cassette, and the essential region for Dunaliella transformation was amplified by PCR and used for transformation. The transformation efficiency using a 3.2kb PCR product was 130 transformants/μg DNA for both sense and antisense transformations. Among 200 BASTA-resistant colonies from the sense transformation and antisense transformation, separately, nine positive transformants for sense expression and five positive transformants for the antisense expression were obtained by genomic DNA PCR. The insertion was also verified by genomic Southern analysis. Among five positive sense transformants, one transformant was tested and verified for the overexpression of CBR-GFP fusion protein by Western blot analysis. One of the five antisense transformants showed almost complete inhibition of gene expression under light stress conditions (400μmol photons m−2 s−1) as determined by quantitative RT-PCR and Western blot analysis. Although there was no difference in the growth patterns or photosynthetic parameters between the wild type (including the vector control) and transformants, the zeaxanthin content of the antisense CBR mutant was lowered under light stress conditions. Thus, we show that the sense and antisense RNA technology can be easily and strategically used for the functional analysis of interesting gene in D. tertiolecta. Springer-Verlag Berlin Heidelberg, 2015 Dunaliella transformation nationallicence Antisense expression nationallicence Carotenoid biosynthesis-related protein nationallicence Overexpression nationallicence Zhang Jing Department of Life Science and Research Institute for Natural Sciences, Hanyang University, 133-791, Seoul, Korea aut Jeon Hancheol Department of Life Science and Research Institute for Natural Sciences, Hanyang University, 133-791, Seoul, Korea aut Sim Sang Department of Chemical and Biological Engineering, Korea University, 136-701, Seoul, Korea aut Lee Yew Department of Life Science and Research Institute for Natural Sciences, Hanyang University, 133-791, Seoul, Korea aut Jin EonSeon Department of Life Science and Research Institute for Natural Sciences, Hanyang University, 133-791, Seoul, Korea aut Planta Springer Berlin Heidelberg 242/4(2015-10-01), 1051-1058 0032-0935 242:4<1051 2015 242 425 https://doi.org/10.1007/s00425-015-2369-2 text/html Onlinezugriff via DOI BK010053 XK010053 XK010000 Metadata rights reserved Springer special CC-BY-NC licence nationallicence 1 brief-communication jats NATIONALLICENCE NATIONALLICENCE NL-springer NATIONALLICENCE 856 40 https://doi.org/10.1007/s00425-015-2369-2 text/html Onlinezugriff via DOI NATIONALLICENCE 700 1- Zhang Jing Department of Life Science and Research Institute for Natural Sciences, Hanyang University, 133-791, Seoul, Korea aut NATIONALLICENCE 700 1- Jeon Hancheol Department of Life Science and Research Institute for Natural Sciences, Hanyang University, 133-791, Seoul, Korea aut NATIONALLICENCE 700 1- Sim Sang Department of Chemical and Biological Engineering, Korea University, 136-701, Seoul, Korea aut NATIONALLICENCE 700 1- Lee Yew Department of Life Science and Research Institute for Natural Sciences, Hanyang University, 133-791, Seoul, Korea aut NATIONALLICENCE 700 1- Jin EonSeon Department of Life Science and Research Institute for Natural Sciences, Hanyang University, 133-791, Seoul, Korea aut NATIONALLICENCE 773 0- Planta Springer Berlin Heidelberg 242/4(2015-10-01), 1051-1058 0032-0935 242:4<1051 2015 242 425