Department of Molecular Biotechnology, Konkuk University, Seoul, South Korea.
Biotechnol Lett. 2010 Feb;32(2):321-30. doi: 10.1007/s10529-009-0140-0. Epub 2009 Oct 11.
L-ascorbic acid (Vitamin C, AsA) is an important component of human nutrition. Plants and several animals can synthesize their own ascorbic acid, whereas humans lack the gene essential for ascorbic acid biosynthesis and must acquire from their diet. In the present study, we developed transgenic potato (Solanum tuberosum L. cv. Taedong Valley) over-expressing L-gulono-gamma-lactone oxidase (GLOase gene; NCBI Acc. No. NM022220), isolated from rat cells driven by CaMV35S constitutive promoter that showed enhanced AsA accumulation. Molecular analyses of four independent transgenic lines performed by PCR, Southern and RT-PCR revealed the stable integration of the transgene in the progeny. The transformation frequency was ca. 7.5% and the time required for the generation of transgenic plants was 6-7 weeks. Transgenic tubers showed significantly enhanced AsA content (141%) and GLOase activity as compared to untransformed tubers. These transgenics were also found to withstand various abiotic stresses caused by Methyl Viologen (MV), NaCl or mannitol, respectively. The T(1) transgenic plants exposed to salt stress (100 mM NaCl) survived better with increased shoot and root length when compared to untransformed plants. The elevated level of AsA accumulation in transgenics was directly correlated with their ability to withstand abiotic stresses. These results further demonstrated that the overexpression of GLOase gene enhanced basal levels of AsA in potato tubers and also the transgenics showed better survival under various abiotic stresses.
L-抗坏血酸（维生素 C，AsA）是人类营养的重要组成部分。植物和几种动物可以合成自己的抗坏血酸，而人类缺乏合成抗坏血酸所必需的基因，必须从饮食中获取。在本研究中，我们开发了过表达 L-古洛糖酸-γ-内酯氧化酶（GLOase 基因；NCBI Acc. No. NM022220）的转基因马铃薯（Solanum tuberosum L. cv. Taedong Valley），该基因来自大鼠细胞，由 CaMV35S 组成型启动子驱动，表现出增强的抗坏血酸积累。通过 PCR、Southern 和 RT-PCR 对四个独立的转基因系进行的分子分析表明，转基因在后代中稳定整合。转化频率约为 7.5%，产生转基因植物所需的时间为 6-7 周。与未转化的块茎相比，转基因块茎表现出明显增强的抗坏血酸含量（141%）和 GLOase 活性。这些转基因植物还被发现能够耐受由甲基紫精（MV）、NaCl 或甘露醇分别引起的各种非生物胁迫。与未转化的植物相比，暴露于盐胁迫（100 mM NaCl）下的 T(1)转基因植物具有更高的成活率，表现出更长的茎和根。转基因植物中抗坏血酸积累水平的升高与它们耐受非生物胁迫的能力直接相关。这些结果进一步表明，GLOase 基因的过表达增强了马铃薯块茎中抗坏血酸的基础水平，并且转基因植物在各种非生物胁迫下表现出更好的生存能力。
