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  本文選題：轉(zhuǎn)基因小麥　切入點：WYMV-Nib8基因　出處：《南京師范大學(xué)》2015年碩士論文

【摘要】：隨著生物技術(shù)的日益發(fā)展和廣泛應(yīng)用,轉(zhuǎn)基因技術(shù)已成為當(dāng)代農(nóng)作物育種的一種重要方法。近年來,轉(zhuǎn)基因作物的商業(yè)化種植面積不斷擴大,隨之而來的擔(dān)憂和社會爭議也越來越多。因此,建立一套科學(xué)、可靠的轉(zhuǎn)基因作物安全評價體系是很有必要的。轉(zhuǎn)基因小麥N12-1轉(zhuǎn)入了小麥黃花葉病毒的復(fù)制酶基因WYMV-Nib8,對小麥黃花葉病具有顯著的抗性。本研究以轉(zhuǎn)基因小麥N12-1、受體小麥揚麥158以及自然抗小麥黃花葉病小麥寧麥9號(N9)為試驗材料。將N12-I、揚麥158于2013-2014年度種植于江蘇省農(nóng)業(yè)科學(xué)院轉(zhuǎn)基因小麥實驗基地(江蘇六合),并于同年將N12-1、揚麥158以及寧麥9號種植于揚州里下河地區(qū)農(nóng)業(yè)科學(xué)研究所轉(zhuǎn)基因小麥種植基地(江蘇揚州)。在小麥生長的播種期、苗期、返青期、抽穗期、灌漿期及成熟期采集小麥根際土壤樣品,并對其微生物生態(tài)、微生物數(shù)量及土壤酶活性進行研究。通過比較轉(zhuǎn)基因小麥N12-1與對照小麥之間的差異性,評價轉(zhuǎn)基因小麥N12-1對其根際土壤生態(tài)系統(tǒng)的影響。研究結(jié)果如下：首先,通過聚合酶鏈?zhǔn)椒磻?yīng)-變性梯度凝膠電泳(PCR-DGGE)技術(shù),對小麥根際土壤中的微生物生態(tài)結(jié)構(gòu)進行分析。結(jié)果顯示,不同生育期小麥根際土壤中細菌和真菌的群落結(jié)構(gòu)差異顯著；另外,不同的生長地區(qū),小麥根際土壤中細菌和真菌群落結(jié)構(gòu)也不相同；然而,種植于六合及揚州地區(qū)的轉(zhuǎn)基因小麥N12-1與對照小麥根際土壤中細菌及真菌的群落結(jié)構(gòu)在各生育期都沒有顯著的差異。然后,利用實時熒光定量PCR (Quantitive Real-Time PCRqPCR)技術(shù),對種植于六合及揚州地區(qū)的轉(zhuǎn)基因小麥N12-1及非轉(zhuǎn)基因小麥根際土壤中熒光假單胞菌(Pseudomonas fluorescens)、鐮刀菌(Fusarium graminearum)和禾谷多黏菌(Polymyxa graminis)的數(shù)量進行檢測。結(jié)果顯示,不同生育期及不同地區(qū)的小麥,其根際土壤中的熒光假單胞菌、鐮刀菌和禾谷多黏菌的數(shù)量均有所不同；但轉(zhuǎn)基因小麥N12-1與對照小麥在各生育期差異都不顯著。最后,分別通過氯代三苯基四氮唑(TIC)法、苯酚-次氯酸鈉比色法和DNS(3,5-二硝基水楊酸)比色法對土壤樣品的脫氫酶活性、脲酶活性及蔗糖酶活性進行測定。結(jié)果顯示,轉(zhuǎn)基因小麥N12-1和對照小麥根際土壤中脫氫酶活性、脲酶活性及蔗糖酶活性隨著生長階段的不同而變化；但是,無論是種植于六合還是揚州,轉(zhuǎn)基因小麥N12-1與其對照小麥根際土壤中脫氫酶活性、脲酶活性及蔗糖酶活性在各生育期均沒有發(fā)現(xiàn)顯著差異。以上研究結(jié)果初步表明,2013-2014年度,種植于江蘇六合及揚州地區(qū)的轉(zhuǎn)WYMV-Nib8基因抗黃花葉病小麥N12-1對其根際土壤中細菌和真菌群落的結(jié)構(gòu),熒光假單胞菌、鐮刀菌和禾谷多黏菌的數(shù)量,以及脫氫酶活性、脲酶活性和蔗糖酶活性均沒有顯著影響。本試驗數(shù)據(jù)為研究轉(zhuǎn)基因小麥N12-1的種植對土壤生態(tài)系統(tǒng)的影響提供了一定的數(shù)據(jù)和技術(shù)參考。
[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.
【學(xué)位授予單位】：南京師范大學(xué)
【學(xué)位級別】：碩士
【學(xué)位授予年份】：2015
【分類號】：S512.1;S154

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