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胚胎发育晚期丰富(LEA)蛋白在植物抵抗非生物胁迫过程中起着重要作用。利用RACE技术从柠条锦鸡儿(Caragana korshinskii)中克隆得到一个LEA_4家族基因Ck LEA4-2,其cDNA全长963 bp,编码240个氨基酸。实时荧光定量PCR检测发现CkLEA4-2的表达受干旱、脱水、NaCl、冷、高pH、脱落酸(ABA)处理的诱导。通过构建35S启动子驱动的pCanG-CkLEA4-2植物过表达载体,研究转基因拟南芥并发现,过表达Ck LEA4-2提高了拟南芥在ABA处理及NaCl和甘露醇胁迫下的萌发率。在幼苗生长阶段,转基因拟南芥的抗旱性增强,失水率低于野生型,并且积累了较少的丙二醛,脯氨酸和谷胱甘肽含量高于野生型。同时,CkLEA4-2过表达后使ABA2、AAO3等ABA信号途径基因以及胁迫响应基因RD22、RD29A和MYB2的表达量升高。结果表明,Ck LEA4-2基因参与了植物逆境应答过程,并提高了转基因拟南芥的抗旱能力。
Late Embryonic Development Rich (LEA) proteins play an important role in plant resistance to abiotic stress. A LEA4 gene Ck LEA4-2 was cloned from Caragana korshinskii using RACE. The full length cDNA of C LEA4-2 is 963 bp, encoding 240 amino acids. Real-time PCR showed that the expression of CkLEA4-2 was induced by drought, dehydration, NaCl, cold, high pH and abscisic acid (ABA). Transgenic Arabidopsis thaliana was constructed by constructing the 35S promoter-driven plant overexpression vector pCanG-CkLEA4-2 and found that overexpression of Ck LEA4-2 increased the Arabidopsis thaliana germination rate under ABA treatment and NaCl and mannitol stress. During the seedling stage, transgenic Arabidopsis plants had higher drought resistance, lower water loss rate than wild type, less accumulated malondialdehyde, and higher contents of proline and glutathione than the wild type. Meanwhile, overexpression of CkLEA4-2 increased ABA2, AAO3 and other ABA signaling pathway genes and the expression of stress response genes RD22, RD29A and MYB2. The results showed that the Ck LEA4-2 gene was involved in plant stress response and increased the drought tolerance of transgenic Arabidopsis thaliana.