Expression And Cloning Of Plasma Membrane Aquaporin Gene From Cucumber Leaf

Drought is a widespread phenomenon of natural disasters. It not only inhibits the growth of crops,but also seriously affects the crops yield and quality. Cucumis sativus L., as an important vegetable cropin our country, plays an important role in Chinese agricultural economy. However, cucumber is underdrought stress when the soil moisture content is less than70%, Because of the cucumber being moresensitive to drought stress, it has a very important significance to cultivate drought-resistant cucumbervarieties by using genetic engineering.Aquaporins are the main plant moisture transmembrane transport channel. Drought is one of theimportant environmental factors limiting the growth and geographical distribution of plant. Despitethere are some relevant reports on the link between drought and aquaporins, the precise mechanism isstill not clear at present. Firstly by building system evolutionary tree, this study identify eight cucumberPIPs with high homology of the arabidopsis thaliana PIPs response to drought stress. In this study,Jinyou No.1is experimental material. By adding PEG6000, simulating nutrient solution culturemethods on drought stress, the research used the real-time PCR to study the relationship betweendrought resistance and aquaporin gene. The study cloned Csa019689aquaporins gene from cucumberby RT-PCR method and constructed eukaryotic expression vector pCAMBIA-Csa019689. Bytransforming arabidopsis plants to obtain arabidopsis seeds. In order to research the cucumber PIPsaffect of drought resistance and the regulatory mechanism of plants further. The main researchs are asfollows:(1) Through analysis on the biological characteristics, we clear eight cucumber PIP genes with thearabidopsis homologous PIPs responsed to drought stress.(2) By adding PEG6000simulating drought stress, setting up the concentration of5%and10%,compared with normal hydroponic plant, in12h,24h and36h three points to collect samples blade. theresult of fluorescence quantitative analysis on cucumber Csa011928, Csa011929, Csa015178,Csa020563, Csa012726, Csa019687, Csa019688, Csa019689showed that Csa019688, Csa019689,Csa011928, Csa011929, Csa020563five genes expression changes Notably, while Csa012726,Csa019687, Csa015178three genes expression changes were not Notably. indicating that PIP1and PIP2subfamily genes had expression of differences when being suffered drought-induced, as well as wasable to response to the stress from the external environment. (3) Having Cloned one Cucumber aquaporin gene and The gene database accession number isCsa019689, belonging to PIP1-4. The full-length of Csa019689cDNA is879bp, encoding292aminoacids. By comparing gene sequence homology, we found that genetic similarity mainly include rubbertrees, cassava, rhubarb, tobacco, arabidopsis.(4)Csa019689gene expression vector was constructed and named pCAMBIA-Csa019689. Vectorwas transformed into Agrobacterium GV3101by freeze-thaw method, and then by Agrobacterium-mediated transformation into Arabidopsis.