过表达天山雪莲SikCDPK1基因提高转基因烟草耐低温能力的机制初探

摘要

CDPKs(calcium-dependent protein kinases) play a very important role in abiotic stress response.Real-time PCR was used to analyze expression of SikCDPK1 gene to cold stress in Saussurea involucrata.Results showed that endogenous SikCDPK1 gene was induced by cold stress,and SikCDPK1 expression reached the peak after 3 h.The SikCDPK1 gene was constructed on dicotyledonous transformation vector pCAMBIA2300,which was composed of a strong promoter of CaMV35S,and the gene was introduced into tobacco NC89 by Agrobacturium-mediated transformation method to identify the gene function.It turned out that after cold stress,cold resistance of transgenic tobacco plants was significantly enhanced,and the content of proline,solubleprotein and the activities of POD and SOD in transgenic tobacco plants were higher than that of wild type,whereas the MDA content and electrolyte leakage were lower than that of wild type plants.These results indicated that SikCDPK1 gene could enhance the resistance of plants to cold stress by accumulating more osmotic adjustment substances,enhancing the activity of the antioxidant system and maintaining the stability of the cell membrane.%钙依赖型蛋白激酶(calcium-dependent protein kinases,CDPKs)在植物参与非生物胁迫信号响应中发挥着重要作用.本研究利用实时荧光定量PCR技术分析了天山雪莲钙依赖性蛋白激酶基因SikCDPK1在-4℃低温胁迫处理下的表达量,发现低温胁迫可以诱导天山雪莲内源SikCDPK1基因表达,且在处理3h表达量迅速积累达到最高值.将SikCDPK1基因构建在由组成型CaMV35S强启动子控制的双子叶转化载体pCAMBIA2300上,利用农杆菌介导法转化烟草NC89,发现转基因烟草植株在低温胁迫后,耐低温能力明显增强;脯氨酸、可溶性蛋白含量及超氧化物歧化酶、过氧化物酶活性均高于非转基因烟草植株;丙二醛含量和相对电导率均低于非转基因烟草植株.上述研究结果表明,SikCDPK1基因可以通过积累渗透调节物质、增强抗氧化酶活性和维持细胞膜稳定性来提高植物的耐低温能力.