【作者】 段苏然；

【导师】 王俊儒； 施卫明；

【作者基本信息】 西北农林科技大学 ， 植物学， 2005， 硕士

【摘要】 随着工业技术的发展,重金属在土壤中的含量越来越高。植物修复可部分解决这一问题且正引起人们的普遍关注。但现在发现许多用于修复的超量积累植物生长缓慢、植株矮小、地上部生物量小,成了实际应用中的最大限制。利用基因工程手段改变植物对重金属吸收、转运、积累和忍耐的机制,从而提高植物对重金属的富集能力,将成为今后植物修复领域研究的一个重要方向。本研究以导入并过量表达了编码大麦pAPX 的基因HvAPX1 的水稻为材料,探讨转基因植物对镉、锌胁迫的耐性提高与否,提高程度以及可能的耐性机制,研究结果为提高植物修复效率提供科学依据。取得的主要结果如下: 1. 用PCR 技术鉴定HvAPX1 基因导入T2 代水稻植株,结果表明,转基因植株的基因组DNA 含有HvAPX1 基因片段,而非转基因水稻植株没有该基因片段。使用RT-PCR 技术验证了导入的HvAPX1 基因在T3 代水稻植株中高量表达。2. 转pAPX 水稻受镉胁迫的实验结果表明,由于pAPX 基因的过量表达,Cd20μM 处理六天后转基因植株叶片内的APX活性始终保持在较高水平,而野生型水稻植株叶片内的APX活性在胁迫初期略微上升后呈现下降趋势;镉胁迫下虽然植株根系的伸长都受到了显著的抑制,但是转基因株系的根系生长受镉的抑制明显低于镉对野生型植株的抑制,表现出一定的耐性;在给镉培养6 天之后,转基因植株和野生型植株间的生物量和叶绿素含量也存在显著差异;Cd20μM 处理六天后,尽管转基因及野生型水稻无论是在根部还是在地上部分,单位干重内的镉含量并没有差异,但是,由于转基因植株生物量明显高于非转基因植株,导致转pAPX 基因水稻在镉胁迫下能够累积更多的镉。3. 转pAPX 水稻受锌胁迫的实验结果表明,由于pAPX 基因的过量表达,转基因植株的根系伸长量、叶绿素含量以及APX 活性都明显高于野生型植株。与野生型植株相比,转基因水稻对锌胁迫具有一定的耐性。但是,与镉胁迫下转pAPX 基因水稻能够累积更多的镉不同的是,转pAPX 基因水稻在锌胁迫下并不能累积更多的锌。更多还原

【Abstract】 With the development of modern industry, heavy metal concentrations in soils have increased unusually. Phytoremediation approach using hyperaccumulator plants, may offer a valuable solution to this problem, and the approach is gaining more attention. But some hyperaccumulators used to remedy the contaminated environment are not ideal for phytoremediation, since they grow slowly and have a low biomass production. These shortcomings limit their practical application. Improving plants tolerance to heavy metals by genetic engineering, i.e. by modifying processes like heavy metal uptake, transport and accumulation opens up new possibilities for phytoremediation. Experiments were conducted to study the effect of overexpressing peroxisomal type ascorbate peroxidase in rice on enhanced tolerance to cadmium and zinc stress. In addition, the mechanism of tolerance to the heavy metals is also studied in this research. The results can offer scientific basis to raise phytoremediation efficiency. The results summarized are as follows: 1. Identification of HvAPX1 in transgenic plants of T2 generation by PCR. The result indicates that the transgenic plants contain the genetic fragment of HvAPX1, but the wide type plants do not have this genetic fragment. Confirmation of transgene HvAPX1 transcript level in the transgenic plants by RT-PCR technology, the result shows that HvAPX1 gene is overexpressed in T3 generation. 2. The results indicated that with the increase of HvAPX1-transcript in transgenic rice, the total APX activity of transgenic plants of both lines kept at high level while the activity decreased obviously in wide type plants. APX activities of HvAPX1-transgenic plants differ significantly from WT after Cd20μM treatment for 6 days. The elongation rate of main root, the total fresh weight and the leaf chlorophyll content are all significantly higher than that of wild type. The Cd content in both root and shoot of transgenic lines show no significantly difference from WT, but the transgenic plants can accumulate more cadmium compared with the wide-type plants. 3. Adding excessive zinc on transgenic and wide type rice showed that with the overexpressing of the pAPX, the elongation rate of the main root, the total fresh weight, the leaf chlorophyll content and the activity of the APX are all higher obviously than that of wild type. The transgenic plants were significantly more tolerant to zinc stress as compared with the wide-type plants. Different from cadmium stress, transgenic plant can not accumulate more zinc.更多还原