慈竹BeCesA基因表达模式分析及其遗传转化毛白杨研究

发布时间：2018-04-03 23:33

  本文选题：慈竹　切入点：毛白杨　出处：《西南科技大学》2017年硕士论文

【摘要】：慈竹(Bambusa emeiensis)是我国西南地区栽培最普遍的竹种之一,其纤维含量较高,是良好的造纸材料。CesA是合成纤维素的关键基因,不仅与纤维素含量相关,也与植物生长发育相关。但有关慈竹Be CesA基因对慈竹生长发育及纤维素含量影响的研究还未见报道,鉴于此,本研究以慈竹笋为材料,克隆获得2条调控纤维素生物合成的BeCesAs基因,对其进行生物信息学分析、表达模式分析和遗传转化研究,为通过基因工程手段改良慈竹提供理论和实践依据。主要研究结果如下:1.在实验室已经得到4条BeCesAs基因全长序列(BeCesA1、BeCesA2、BeCesA7和BeCesA8)的基础上,克隆获得2条慈竹BeCesAs基因序列,分别命名为BeCesA3和BeCesA4,其编码序列长度分别为3282bp和2949bp,分别编码1093和982个氨基酸残基。GenBank注册号分别为KX424591和KX424592。2.生物信息学分析结果表明,BeCesA1、BeCesA2、BeCesA3、BeCesA4、BeCesA7和BeCesA8分别与OsCesA1、BoCesA3、OsCesA3、OsCesA4、OsCesA7和BoCesA4的亲缘关系最近。六条慈竹BeCesA1,2,3,4,7,8蛋白序列拥有所有CesA蛋白及糖转运蛋白都具有的4个保守基序(D,DxD,D,Q/RxxRW),一段植物CesA高度保守区域(P-CR),两个高度可变区域(VR1和VR2),一段酸性氨基酸富集区域(ARR)。3.组织表达模式分析结果表明,BeCesA1,3在未展开叶和30cm笋中表达量较高,在展开叶和100cm笋中表达量较低;BeCesA4,7与BeCesA1,3的表达模式相反。进一步测定30cm、50cm和170cm笋中BeCesA1,3,4,7基因的相对表达水平发现,随着慈竹笋高度增加,BeCesA1,3基因的相对表达水平逐渐下调,而BeCesA4,7基因的表达趋势与BeCes A1,3基因完全相反。4.对慈竹实生植株进行GA_3和DMZ诱导处理。结果表明,在未展开叶中,GA_3和DMZ都能促进BeCesA3,4的表达,而在展开叶中,GA_3会促进BeCesA3表达,而抑制BeCesA4表达。GA_3通过促进细胞伸长使得慈竹侧枝节间伸长,而DMZ通过促进细胞分裂使得节间伸长。但两者对慈竹纤维素含量的影响却不同,GA_3处理后慈竹茎秆中纤维素含量增加,展开叶中含量降低;DMZ处理后慈竹茎秆及叶片中纤维素含量都降低。进一步研究发现,慈竹NAC29/31、MYB61与BeCesA的表达呈极显著正相关关系。5.构建了BeCesA3,4基因的植物过表达载体pCAMBIA1303-N-BeCesA3,4,并用农杆菌介导重组质粒pCAMBIA1303-N-BeCesA4遗传转化毛白杨,得到9株转基因阳性植株,其中只有7株转基因阳性植株的BeCesA4表达量较高,其株高、叶片长度和叶片宽度均显著高于对照,但叶片纤维素含量显著低于对照。
[Abstract]:Bambusa emeiensis is one of the most common bamboo species cultivated in southwest China, and its fiber content is high. CesA is a good paper-making material, which is the key gene of cellulose synthesis, which is not only related to cellulose content, but also related to plant growth and development.However, the effect of be CesA gene on the growth, development and cellulose content of Cesicarachys sinensis was not reported. In view of this, two BeCesAs genes regulating cellulose biosynthesis were cloned from the bamboo shoots.The bioinformatics analysis, expression pattern analysis and genetic transformation study provide theoretical and practical basis for improving the Zizhiya chinensis by means of genetic engineering.The main results are as follows: 1.On the basis of four full-length sequences of BeCesAs gene BeCesA1, BeCesA2, BeCesA7 and BeCesA8, two BeCesAs genes were cloned.They were named BeCesA3 and BeCesA4, respectively. The length of coding sequence was 3282bp and 2949bprespectively, encoding 1093 amino acid residues and 982amino acid residues respectively. GenBank registration numbers were KX424591 and KX424592.2, respectively.鐢熺墿淇℃伅瀛﹀垎鏋愮粨鏋滆〃鏄，

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