日本结缕草 ZjADH 基因对拟南芥的转化及其耐寒性分析

为研究日本结缕草ZjADH基因是否与植物耐寒有关,通过DNA重组技术将ZjADH基因插入3302Y质粒中,成功构建了植物表达载体3302Y-ZjADH,通过农杆菌介导的花序侵染法获得具有草铵膦抗性的转基因植株。PCR和qRT-PCR鉴定表明,目的基因已整合入拟南芥基因组中并能够成功表达。对野生型和转基因拟南芥进行低温(4℃)处理和耐寒分析,结果表明,转基因拟南芥对低温胁迫的抵抗能力明显高于未转基因植株。在低温胁迫下,转基因拟南芥中脯氨酸的含量显著高于野生型植株,而活性氧的积累明显低于野生型植株;对转基因植物抗性相关基因的表达分析显示:转基因植物体内SOD、APX及LEA基因的表达水平明显高于未转基因植株,说明ZjADH基因可能通过提高转基因植物体内SOD、APX及LEA的表达活性来增强植物的抗寒性。综上所述,日本结缕草ZjADH基因的表达能够提高转基因植株的耐寒性,对ZjADH基因的研究也为进一步获得抗寒转基因日本结缕草植株奠定理论依据。

Transformation of ZjADH gene into Arabidopsis thaliana and cold-tolerance analysis of transgenic plants

To study the relationship between the ZjADH gene and plant cold tolerance,a plant expression vec-tor 3302Y-ZjADH was constructed through DNA recombination technology.Transgenic Arabidopsis thaliana plants with glufosiante resistance were generated by floral dip method.PCR and qRT-PCR results showed that the target gene was successfully integrated with the genomes of Arabidopsis and could express efficiently. Both wild type and transgenic Arabidopsis plants were subjected to 4 ℃ to investigate their cold tolerance. Tests demonstrated that,compared with the wild type,the transgenic plants were more tolerant of cold stress. The free proline content in transgenic plants was significantly higher than the control,while the accumulation of ROS was much lower than the wild type.In addition,the expression levels of genes related to stresses like SOD ,APX and LEA in transgenic plants were much higher than in the control.Results indicated that over-ex-pression of ZjADH may improve the activities of superoxide dismutase (SOD),ascorbate peroxidase (APX) and late embryogenesis abundant proteins (LEA)to improve the cold tolerance of transgenic plants.In conclu-sion,the heterologous expression of ZjADH genes could reinforce the resistance of plants to cold stress.This study provides a theoretical foundation for future research on the ZjADH gene and the cultivation of cold-toler-ant Zoysia japonica cultivars.