关键词: 杨树, Rubisco活化酶基因, 转基因, 光合作用

Abstract: [Objective] Ribulose-1,5-bishosphate (Rubisco) is the key enzymes in the first step of carbon assimilation involved in plant photosynthesis. Rubisco was kept in a catalytic active state under the control of Rubisco activase(RCA), as a result, the efficiency of photosynthesis will be improved. In this research, the PtRCA gene of poplars was cloned. Transgenic poplars with high expression of PtRCA were obtained through this research. As a result, a new type of high efficiency photosynthesis with stress tolerance was proved to be feasible through the analysis of molecular detection and functional analysis.[Method] According to the sequence of PtRCA gene cloned from Populus trichocarpa, the analysis of function and structure of PtRCA were conducted through Bioinformatics software. PtRCA was constructed into the expression vector pGWB406 by using Agrobacterium mediated method. 'Nanlin895'poplar plants (P. deltoides×P. euramericana 'Nanlin895') were used in this research. Gene expression of both transgenic poplars and 'Nanlin895'plants were compared under high temperature stress. The parameters of photosynthesis and chlorophyll fluorescence were compared between them as well.[Result] The CDS sequence of PtRCA was 1 323 bp. There were 440 amino acid residues, and protein molecular weight is 48 315.9 Da. The isoelectric point is 5.57, which is hydrophobic protein without signal peptide and membrane structure. PtRCA is proved to be in the same AAA+ family according to the result of sequence comparison. It has same RCA homology protein with soybean and arabidopsis as well. The expression quantity of transgenic poplars were higher than 'Nanlin895' in average. Transgenic poplars showed advantage of photosynthesis during the noon, the light saturation point was 12.5% to 37.5% higher. Moreover, the efficiency of photosynthesis was 24.6% to 55.7% higher than the plants used as control. However, the light compensation point is lower than that of plants without transgenosis expect the No.3 strain. Besides, the usage of CO₂ and the carboxylation efficiency of the transgenic poplars were better. Expect No.1 and No.4 strains, CO₂ saturation point was around 12.5%-25.0% less than control plants. CO₂ compensation point and light respiration were proved to be 53.1%-80.4% and 37.7%-79.3% lower compared to the controls respectively. Moreover, the efficiency of photosynthesis at the CO₂ saturation point was around 4.4%-26.4% higher. The tolerance of transgenic poplars to light oxidation was increased according to this research. Under the light oxidation treatment, the primary photochemical efficiency of PSⅡ decreased 61.7% in control lines. And transgenic poplar decreased by 45.0%- 53.1%.The actual photochemical efficiency of PSⅡ in controls decreased 54.1%. And transgenic poplar decreased by 38.7%-52.0%.Photochemical quenching coefficient dropped 68.3%, and 51.0%-65.8% declined in transgenic poplars. Non photochemical quenching coefficient increased by 3.0% compared to the control, and 6.0%-26.5% increased in transgenic poplars.[Conclusion] Poplar Rubisco activase (PtRCA) protein and soy protein, Arabidopsis thaliana RCA homology is higher. According to the real-time quantitative PCR and related physiological analysis, transgenic poplars (P. deltoides×P. euramericana 'Nanlin895') showed high-temperature resistance, higher efficiency of photosynthesis, increased ability to use CO₂ and better catalyzing effect on carboxylation reaction. The efficiency of assimilating photons and supplying to PSⅡ was highly developed, and as a result, the light oxidation ability of transgenic poplars was proved to be enhanced. The efficiency of photosynthesis was improved by high expression of PtRAC gene in transgenic poplars, which also showed ability to adjust for high temperature and strong light.

Key words: Populus, Rubisco activase gene, transgenic, photosynthesis