The Generation And Research Of Calnexin-antisense Transgenic Tomato Plant

Lectin-like chaperone proteins in ER are mostly Calnexin and Calreticulin. Calnexin is a type I membrane protein, which can assist glycoproteins with N-linked oligosaccharide to fold and assembly. Calnexin plays an important role in ER quality control, degrading unfolded proteins, modulating Ca2+ homeostasis of ER and apoptosis. The calnexin gene was distributed widely in animals, plants and microorganism. Furthermore, the structure and sequence of the calnexin genes was conservatism in various species by sequence analysis. Calnexin plays important roles in animal cells, but the function of Calnexin in plant cells has been less researched. A cDNA encoding a full-length calnexin was cloned from Lycopersicon esculentum previously, and designated as Lecnx61.0 (AB218598). Previous studies showed that the expression of Lecnx61.0 was induced by heat, cold and salty stress, but had no response to drought stress.In this study, an antisense Lecnx61.0 cDNA fragment was introduced into tomato by Agrobacterium-mediated transformation. The results showed that the resistant ability to salt stress of tansgenic plants was similar to that of control plants. The results were shown as follows:1. The generation of tomatoes with repressed Lecnx61.0In order to study the function of Lecnx61.0 in stress conditions, we introduced an antisense Lecnx61.0 cDNA fragment into the tomato by Agrobacterium-mediated transformation. PCR analysis showed that foreign gene had been introduced into tomato genome. Western blotting analysis showed that the expression of the Lecnx61.0 protein was repressed.2. Analysis on resistance to salt stress of transgenic tomatoThe transgenic plants and controls were treated by 150 mmol/L NaCl. The plant growth was obviously repressed after salt treatment for 30 days, and partial leaves scorched. The growth state of the transgenic plants had no obvious difference from the controls. A set of parameters(relative water content, Na+ , K+ content, proline content, soluble sugar content, MDA content, relative conductivity, SOD enzyme activity, chlorophyl content)were measured in transgenic plants and the controls after the treatment. The result was that these parameters had no difference between the transgenic plants and the controls.Comparative analysis of the Lecnx61.0 transgenic tomatoes and non-transgenic tomatoes on salt stress treatment indicated that anti-expressing Lecnx61.0 in plants had no effect on their salt stress resistance ability.