CAJ | 학술논문

构建了同时含有胞质谷氨酰胺合成酶(GS1)cDNA和叶绿体谷氨酰胺合成酶(GS2)cDNA的植物表达载体p2GS,通过农杆菌介导法用它们转化了水稻品种"中花10号"的成熟胚愈伤组织,经潮霉素(Hyg)筛选培养及分化再生,获得了抗Hyg的转基因水稻植株.PCR和基因组Southern杂交分析结果证明,GS1和GS2基因均已经整合到转基因水稻的基因组内.Northern杂交实验结果证实,GS1和GS2基因在转基因水稻的转录水平上得到了有效表达.在以0.7 mmol/L的(NH4)2SO4取代了其中氮成分的MS培养基上测试植株生长量,结果表明转基因植株鲜重增长量显著高于对照,证明高效表达GS增强了转基因水稻对土壤氮素缺乏的耐性.
구건료동시함유포질곡안선알합성매(GS1)cDNA화협록체곡안선알합성매(GS2)cDNA적식물표체재체p2GS,통과농간균개도법용타문전화료수도품충"중화10호"적성숙배유상조직,경조매소(Hyg)사선배양급분화재생,획득료항Hyg적전기인수도식주.PCR화기인조Southern잡교분석결과증명,GS1화GS2기인균이경정합도전기인수도적기인조내.Northern잡교실험결과증실,GS1화GS2기인재전기인수도적전록수평상득도료유효표체.재이0.7 mmol/L적(NH4)2SO4취대료기중담성분적MS배양기상측시식주생장량,결과표명전기인식주선중증장량현저고우대조,증명고효표체GS증강료전기인수도대토양담소결핍적내성.
Glutamine synthetase (GS, EC6.3.1.2) is a key enzyme in ammonia assimilation both in plants and in Gram-negative microorganisms. It plays an important role in efficient use of nitrogen sources and nitrogen metabolism in organisms. Two groups of GS isoenzymes, plastidic (GS2) and cytosolic (GS 1), have been identified in higher plants. A plant constitutive expression vector p2GS harboring GS1 and GS2 under the control of rice actin 1 (Actl) and maize ubiquitin (Ubi) promoters was constructed for the first time in a single plasmid, and 3 rounds of ligation and transformation were performed. There has been no report about studies on rice transformation with the two GS enzymes (GS1 and GS2). The p2GS thus constructed was introduced into rice var. Zhonghua 10 by Agrobacterium-mediated transfer method, and transgenic plants with resistance to hygromycin (Hyg)were obtained. Results of PCR and Southern blot analysis showed that the foreign genes have been integrated into the rice genome. The transcription of GS1-GS2 genes in the transformants was also confirmed by Northern blot analysis. The transgenic rice plants thus obtained can grow well in an MS medium in which the nitrogen source was replaced by (NH4)2SO40.7mmol/L, and fresh weight of the transformants was significantly higher than the control rice plants. The result suggests that expression of p2GS makes the transgenic rice plants tolerant to nitrogen-deficiency.