【作者】 胡亚杰；

【导师】 刘卫群；

【作者基本信息】 河南农业大学 ， 烟草学， 2008， 硕士

【摘要】 水分是保证烟草健康生长发育、优良品质形成的主要因素之一。如果干旱出现在团棵期,易造成烟株早花;出现在旺长期,会造成叶片发育不良、内含物不充实;出现在成熟期,会导致叶片增厚、粗糙,糖碱比失调,旱烘假熟现象。DREB(dehydration responsive element binding protein)类转录因子主要参与植物感受非生物逆境胁迫的信号传递途径,通过调控下游一系列胁迫应答基因的表达,使植物对低温、干旱和高盐等非生物逆境的耐受性得到综合提高。有关DREB类转录因子的结构、功能及对效应基因的调控研究已有相当多的报道,但DREB类转录因子过表达对植物生物学特性及对渗透胁迫耐受性的生理过程的综合研究却少见报道。本研究就香料烟(Izmir)中过表达BnDREB1-5转录因子基因对叶片组织结构、光合特性的影响及抗胁迫能力进行了初步探讨。主要结果如下:1.转基因香料烟表现出良好的保水性能,叶片失水率为0.16 mg H2O / cm2 .h-1,是野生型烟草叶片的15.9%;转基因烟草叶片上表皮的气孔密度是野生型叶片气孔密度的46.4%,但气孔器宽度和气孔开度都略大于野生型;下表皮是野生型的56.1%,但气孔器长于野生型,宽微小于野生型,气孔开度明显大于野生型。2.检测转基因烟株在干旱胁迫10d后的农艺性状比野生型烟草有明显优势,(T/W单位cm:株高51.67/39.67、叶数17.67/13.67、叶长19.95/19.22、叶宽9.93/10.61、节距2.97/2.98),且野生型烟株叶片明显萎焉;组织结构分析显示:转基因烟草叶片增厚,栅栏组织/海绵组织的比值明显增大,组织结构完整。3.干旱胁迫后转基因烟叶中淀粉酶活性是野生型的1.20倍;Ivr活性是野生型的1.71倍;可溶性糖含量是野生型的1.86倍。NR活性是野生型的8.47倍;可溶性蛋白质是野生型的2.97倍。4.干旱胁迫后转基因烟叶中叶绿素含量降低幅度明显小于野生型;类胡萝卜素和chla/b明显大于野生型烟草;叶绿素荧光参数Fo、NPQ明显低于野生型,其它荧光参数和光合作用参数都大于野生型;PEP羧化酶活性明显大于野生型。5.转基因烟叶中有BADH活性,干旱胁迫后BADH活性升高,而野生型烟草中检测不到BADH活性;干旱胁迫后转基因烟叶中脯氨酸含量是野生型的1.08倍; SOD活性明显高于野生型;MDA含量是野生型的53.9%。以上结果表明:DREB类转录因子过表达烟草提高了PEP羧化酶活性,显现了BADH的活性,并且在保水性能、农艺性状、光合特性及防御酶活性等方面明显优于野生型烟草,DREB类转录因子过表达提高了烟草抗旱的生理机能。更多还原

【Abstract】 Water is one of the main factors to guarantee tobacco healthy growth and development and good quality formation. If drought happened in resettling stage, tobacco plants result in premature flowering. If which happened in vigorous growing stage, the development of leaves is bad, and the inclusion is not rich. If which happened in maturity stage, the leaves are thick and rough, sugar/nicotine ratio is imbalance, the phenomenon of drought cook and premature happen too. DREB (dehydration responsive element-binding protein) transcription factor is mainly involved in signaling pathway of obiotic stresses by plant perception. Which improves the comprehensive tolerance of low-temperature, drought, high-salt and other stresses of plants by regulated the expression of a series of downstream stress response genes. The reports on DREB-like transcription factor structure, function and studies of regulating effect gene are a lot, but there are few reports on biological properties and the physiology process of osmotic stress tolerance of plants over-expressed by DREB-like transcription factor. The tissue structure, photosynthetic characteristics and stress resistance of BnDREB1-5 transcription factor over-expression on oriental tobacco (Izmir) were preliminarily studied. The main results were as follows:1.The leaves of Izmir with BnDREB1-5 presented a better holding water capacity,the water-loss was 0.16 mg H2O / cm2 .h-1 , and was 15.9% of that of wild-types. The upper epidermis stomata density in transgenic tobacco leaves was 46.4% as large as in wild-type leaves, the width of stomata apparatus and stoma opening were slightly bigger than that of wild-type. And the lower epidermis stomata density was 56.1% as large as in wild-type leaves, the length of stomata apparatus was bigger than that of wild-type, the width was slightly smaller than that of wild-type, the stoma opening was obviously bigger than that of wild-type.2.The agricultural characters of transgenic tobacco were obviously better than that of wild-type after drought stress tenth day (T/W unit cm: plant height 51.67/39.67, leaf number 17.67/13.67, leaf length 17.67/13.67, leaf width 9.93/10.61, pitch 2.97/2.98), and the leaves of wild-type plant obviously wilted. The tissue structure analysis showed that the leaf thickness of transgenic tobacco leaves increased, the ratio between palisade and spongy tissue obviously increased, tissue structure was integrate.3. After drought stress, the amylase activity, the invertase activity, the soluble sugar content, the nitrate reductase activity, the soluble protein content of transgenic tobacco leaves were respectively 1.20 times, 1.71 times, 1.86 times, 8.47 times and 2.97 times as large as those of wild-type.4. After drought stress, the chorophyll content descreased degrees of transgenic tobacco leaves were significantly lower than that of wild-type. The carotenoid content and chla/b were obviously bigger than that of wild-type. The fluorescence parameters Fo and NPQ were significantly lower than that of wild-type, the parameters of other fluorescence and photosynthesis were bigger than those of wild-type. The PEP carboxylase activity was obviously bigger than that of wild-type.5. The BADH activity of transgenic tobacco leaves was detectable and increased after drought stress, but which of wild-type was not detectable. After drought stress, the proline content of transgenic tobacco leaves was 1.08 times as large as that of wild-type. The SOD activity was obviously bigger than that of wild-type. The MDA content was 15.9% of that of wild-types.These results showed that by DREB-like transcription factor over-expressed on tobacco, the PEP carboxylase activity increased, the BADH activity was detectable, the holding water capacity, agricultural characters, photosynthetic characteristic and defense enzyme activity and so on were obviously better than those of wild-type. The DREB-like transcription factor overexpression improved tobacco physiological function of drought resistance.更多还原