羊草LcPIP基因遗传转化露地菊及其抗盐性鉴定

Genetic transformation of LcPIP gene in Chrysanthemum morifolium and identification of its salt resistance

PIP是重要的水孔蛋白之一,与植物的抗逆性有关。本文用叶盘法将羊草LcPIP遗传转化露地菊火焰,通过常规PCR和GUS染色方法鉴定转基因株系,Real-time PCR检测盐胁迫下转基因和野生植株中CmPIP1和CmPIP2的相对表达量,并测定SOD、POD活性和MDA含量。结果显示,在转LcPIP基因的露地菊植株叶片中,CmPIP1和CmPIP2基因表达量均上升,是野生型植株的2倍;根部的CmPIP1与CmPIP2基因的表达量均上升,CmPIP1上升程度高于CmPIP2。在200 mmol·L^(-1) NaCl的胁迫下,野生型植株中CmPIP1基因表达量明显下降,而转基因植株中CmPIP1基因表达量在12~48 h出现明显的上升;在盐胁迫12 h后,转基因植株中CmPIP2基因表达量上升程度明显高于野生型植株。盐胁迫12 h后,转基因植株的SOD活性提高更为明显。在盐胁迫前期(0~24 h)转基因植株MDA含量增加幅度低于野生型植株;在盐胁迫后期(48~72 h)转基因露地菊POD活性出现明显上升,而野生型露地菊呈下降趋势。

PIP is one of the important aquaporins, and play a crucial role in plant stress tolerance. We transfered Lc PIP into Chrysanthemum morifolium huoyan by using the leaf disc method, and identified transformed plants by the PCR and GUS staining methods. The relative expression of CmPIP1 and CmPIP2 in transgenic and WT plants under salt stress were determined by real-time PCR, and the activities of SOD, POD and MDA content were also determined. The results showed that the expression of CmPIP1 and CmPIP2 increased in the leaves of transgenic plant, which were two times higher than those of WT plants. The expression of CmPIP1 and CmPIP2 also increased in the roots of transgenic plant, and the rise of CmPIP1 was higher than that of CmPIP2. Under the stress condition of 200 mmol·L^（-1） NaCl, the expression of CmPIP1 decreased obviously in WT plants, while which in the transgenic plants increased significantly at 12–48 h stress. After 12 h salt stress, the increase of CmPIP2 gene expression in transgenic plants was significantly higher than that of WT plants, and the activity of SOD increased significantly. In the early stage of salt stress（0–24 h）, the MDA content of transgenic plants increased less than that of WT plants. The POD activity of transgenic plant increased significantly in the late stage of salt stress（48–72 h）, while that of WT plant showed a decreasing trend.