转WYMV-Nib8基因小麦对其根际土壤微生物生态及主要酶活性的影响

发布时间：2018-04-09 07:39

本文选题：转基因小麦　切入点：WYMV-Nib8基因　出处：《南京师范大学》2015年硕士论文

【摘要】：随着生物技术的日益发展和广泛应用,转基因技术已成为当代农作物育种的一种重要方法。近年来,转基因作物的商业化种植面积不断扩大,随之而来的担忧和社会争议也越来越多。因此,建立一套科学、可靠的转基因作物安全评价体系是很有必要的。转基因小麦N12-1转入了小麦黄花叶病毒的复制酶基因WYMV-Nib8,对小麦黄花叶病具有显著的抗性。本研究以转基因小麦N12-1、受体小麦扬麦158以及自然抗小麦黄花叶病小麦宁麦9号(N9)为试验材料。将N12-I、扬麦158于2013-2014年度种植于江苏省农业科学院转基因小麦实验基地(江苏六合),并于同年将N12-1、扬麦158以及宁麦9号种植于扬州里下河地区农业科学研究所转基因小麦种植基地(江苏扬州)。在小麦生长的播种期、苗期、返青期、抽穗期、灌浆期及成熟期采集小麦根际土壤样品,并对其微生物生态、微生物数量及土壤酶活性进行研究。通过比较转基因小麦N12-1与对照小麦之间的差异性,评价转基因小麦N12-1对其根际土壤生态系统的影响。研究结果如下：首先,通过聚合酶链式反应-变性梯度凝胶电泳(PCR-DGGE)技术,对小麦根际土壤中的微生物生态结构进行分析。结果显示,不同生育期小麦根际土壤中细菌和真菌的群落结构差异显著；另外,不同的生长地区,小麦根际土壤中细菌和真菌群落结构也不相同；然而,种植于六合及扬州地区的转基因小麦N12-1与对照小麦根际土壤中细菌及真菌的群落结构在各生育期都没有显著的差异。然后,利用实时荧光定量PCR (Quantitive Real-Time PCRqPCR)技术,对种植于六合及扬州地区的转基因小麦N12-1及非转基因小麦根际土壤中荧光假单胞菌(Pseudomonas fluorescens)、镰刀菌(Fusarium graminearum)和禾谷多黏菌(Polymyxa graminis)的数量进行检测。结果显示,不同生育期及不同地区的小麦,其根际土壤中的荧光假单胞菌、镰刀菌和禾谷多黏菌的数量均有所不同；但转基因小麦N12-1与对照小麦在各生育期差异都不显著。最后,分别通过氯代三苯基四氮唑(TIC)法、苯酚-次氯酸钠比色法和DNS(3,5-二硝基水杨酸)比色法对土壤样品的脱氢酶活性、脲酶活性及蔗糖酶活性进行测定。结果显示,转基因小麦N12-1和对照小麦根际土壤中脱氢酶活性、脲酶活性及蔗糖酶活性随着生长阶段的不同而变化；但是,无论是种植于六合还是扬州,转基因小麦N12-1与其对照小麦根际土壤中脱氢酶活性、脲酶活性及蔗糖酶活性在各生育期均没有发现显著差异。以上研究结果初步表明,2013-2014年度,种植于江苏六合及扬州地区的转WYMV-Nib8基因抗黄花叶病小麦N12-1对其根际土壤中细菌和真菌群落的结构,荧光假单胞菌、镰刀菌和禾谷多黏菌的数量,以及脱氢酶活性、脲酶活性和蔗糖酶活性均没有显著影响。本试验数据为研究转基因小麦N12-1的种植对土壤生态系统的影响提供了一定的数据和技术参考。
[Abstract]:With the development and wide application of biotechnology, transgenic technology has become an important method of crop breeding.In recent years, the commercial planting area of genetically modified crops has been expanding, and there are more and more worries and social controversies.Therefore, it is necessary to establish a set of scientific and reliable transgenic crop safety evaluation system.Transgenic wheat N12-1 was transformed into WYMV-Nib8, a replicase gene of wheat yellow mosaic virus.In this study, transgenic wheat N12-1, receptor wheat Yangmai 158 and natural resistant wheat Nimai 9 (N9) were used as experimental materials.N12-Iand Yangmai 158 were planted in the experimental base of transgenic wheat of Jiangsu Academy of Agricultural Sciences in 2013-2014. N12-1, Yangmai 158 and Nimai 9 were planted in Yangzhou Lixiahe Research Institute of Agricultural Sciences in the same year.Wheat planting Base (Yangzhou, Jiangsu Province).Wheat rhizosphere soil samples were collected at seeding stage, seedling stage, green stage, heading stage, grain filling stage and maturity stage of wheat growth, and their microbial ecology, microbial quantity and soil enzyme activity were studied.The effects of transgenic wheat N12-1 on rhizosphere soil ecosystem were evaluated by comparing the difference between transgenic wheat N12-1 and control wheat.The results are as follows: firstly, the microbial ecological structure in wheat rhizosphere soil was analyzed by polymerase chain reaction-denaturing gradient gel electrophoresis (PCR-DGGE) technique.The results showed that the community structure of bacteria and fungi in rhizosphere soil of wheat at different growth stages was significantly different, in addition, the community structure of bacteria and fungi in rhizosphere soil of wheat was also different in different growing areas.The community structure of bacteria and fungi in rhizosphere soil of transgenic wheat N12-1 planted in Liuhe and Yangzhou areas was not significantly different from that of control wheat at different growth stages.Then, the quantity of Pseudomonas fluorescens (Pseudomonas fluorescensus) and Polymyxa graminis (Polymyxa graminis) in the rhizosphere soil of transgenic wheat N12-1 and non-transgenic wheat in Liuhe and Yangzhou areas were detected by real-time quantitative PCR Real-Time PCRqPCR.The results showed that the number of Pseudomonas fluorescein, Fusarium and polymyxomycetes in rhizosphere soil of wheat at different growth stages and different regions were different, but there was no significant difference between the transgenic wheat N12-1 and the control wheat at each growth stage.Finally, the dehydrogenase activity, urease activity and sucrase activity of soil samples were determined by triphenyl tetrazolium chloride, phenol-sodium hypochlorite colorimetry and DNSN 3N 5 dinitrosalicylic acid colorimetry respectively.The results showed that dehydrogenase activity, urease activity and sucrase activity in rhizosphere soil of transgenic wheat N12-1 and control wheat changed with different growth stages, but both in Liuhe and Yangzhou,There was no significant difference in dehydrogenase activity, urease activity and sucrase activity in rhizosphere soil between transgenic wheat N12-1 and control wheat.The results indicated that the WYMV-Nib8 gene resistant wheat N12-1 cultivated in Liuhe and Yangzhou area of Jiangsu Province in 2013-2014 showed the structure of bacterial and fungal communities, the number of Pseudomonas fluorescein, Fusarium oxysporum and polymyxomycetes in the rhizosphere of wheat N12-1.Dehydrogenase activity, urease activity and sucrase activity were not significantly affected.The experimental data provided some data and technical reference for studying the effect of transgenic wheat N12-1 on soil ecosystem.
【学位授予单位】：南京师范大学
【学位级别】：硕士
【学位授予年份】：2015
【分类号】：S512.1;S154

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