The Effects Of Overexpression Or Inducible-expression Of APX On Stress Tolerance In Poplar (Populus Tomentosa)

The imposition of environmental stress leads to increased accumulation of reactive oxygen species (ROS) in plant cells, ROS can cause extensive cell injury if they can not be scavenged efficiently. In plants, the scavenging mechanisms of peroxides can be classified into two categories: enzymatic scavengers and nonenzymatic antioxidants. Ascorbate peroxidase (APX) is an heme-binding enzyme that reduce H₂O₂ to water using ascorbate as an electron donor, it is one of the important enzymes of enzymatic detoxication system in plants. Plants contain several APX isoenzymes, and chloroplasts APXs (chlAPXs) have been considered as key enzymes in the detoxification of H₂O₂ in chloroplasts.The cauliflower mosaic virus (CaMV) 35S promoter is one of the constitutive promoters,which is in common use on plant transgenetic engineering. It can promote transcription effectively in nearly all the plant tissues and all the growth phase. Arabidopsis rd29 promoter is a strees inducible promoter, which is only induced by high-salt, cold and desiccation conditions or exogenous ABA.We used Agrobacterium tumefaciens to transform target gene into 741 poplar, to analyze the effects of overexpression or inducible-expression of APX on stress tolerance in 741 poplar. The results were shown as follows:1) PCR and Real-time PCR: The result of PCR showed that the poplar plants transformed with the target gene fragment had been amplified DNA segments with the NPTII primers, while the wild type haven't, which indicate that the pBI121-APX,pBI101-RD29-APX fragment had been transformed into the poplars. The Real-time PCR showed that, under normal growing conditions, the transcription level of APX gene in the 35S-APX lines were higher than control plants, while the inducible-expression ones were as same as the control plants.2) Treatment of methyl viologen(MV): After treated with 0.02mM MV and kept in the plant growth chamber with high light at 28℃for four days, damage was observed in all of the leaf discs, but the damage of control plants was even more serious than that of transgenic ones. Decrease in the content of chlorophyll after treatment with 0.02 mM MV was lower in the transgenic lines than in the control plants.3) Salt stress: 50 mM NaCl nearly restrained the rooting of control poplar in vitro, but the 35S-APX or RD29-APX lines could still root. Interestingly, we found that the RD29-APX lines had suffered more damage, when grew in 25mM NaCl culture medium than in 50mM NaCl ones. Salt tolerance of the potted seedlings was also tested. After treatment with 50 mM NaCl for four days, the activity of APX and SOD in transgenic or control poplar leaves all increased. The content of MDA in plant leaves was also mensured, and the increase of the content of MDA was the highest in control plants.