The Functional Research On CbF15and AtT26J14.6

Chorispora bungeana is a typical alpine periglacial plant species. Chorispora bungeana can grow and develop normally under anaerobic environment, barren soil, and strong radiation, at the high altitude(2600-4200m above sea level) and low temperatures(about0℃), completing its whole life cycle without hindrance by these stress. This indicates strong capability of surviving in an environment with extremely low temperatures. This experiment conducted with Chorispora bungeana as the experimental material. We have selected the Chorispora bungeana gene CbF15, up-regulated in low temperatures screened out by the method of suppression subtractive hybridization, and AtT26J14.6(which has an amino acid sequence homology of80%with CbF15) to research their gene functions by using methods such as RACE, Gateway Cloning, Western-blot, and transient expression of fluorescent fusion proteins in tobacco plants. The results are as follows:1. The whole cDNA sequence of CbF15was cloned by RACE. Furthermore,bioinformatics analysis and structure prediction were conducted. The cDNA length of CbF15is1467bp,containing555bp as an open reading frames which encoded185amino acids with the molecular weight (MW)20.24KD. The main secondary structure of CbF15is random coil. Through a hydrophobicity analysis it could be concluded that CbF15is a transmembrane protein.2. The subcellular localizations of CbF15and AtT26J14.6were identified by transient expression of fluorescent fusion proteins in tobacco.They were both localized on cell membranes.3. Purified proteins and antibodies of CbF15and AtT26J14.6were obtained. By Western-blot it was revealed that CbF15is an early cold-and salt-responsive protein, due to the increase in its early expression under cold and NaCl treatment.4. It was also revealed that the AtT26J14.6gene plays a role in the development process of Arabidopsis thaliana under stress treatment. The AtT26J14.6gene participates in cotyledon development under salt and ABA treatment. Another finding was that AtT26J14.6plays a role in frost resistance of Arabidopsis thaliana.5. It was found that the AtT26J14.6gene expresses in different development stages, and various organs of Arabidopsis thaliana, especially in the meristem with greater expression.6. Analysis of the relationship between CbF15gene and AtT26J14.6gene. CbF15gene has high homology and similar function with AtT26J14.6gene.To sum up, this research has not only added a new member to the Chorispora bungeana Genes Library, but also made a start and foundation of research in the functions of CbF15and AtT26J14.6genes of which were previously unknown. It provided a new theoretical basis for the molecular mechanism of stress resistance and the participation of these two genes in it. As a result new foundations for the exploration of plant resistance gene resources were layed.