转Bt基因玉米的抗螟性及产量分析

在玉米心叶期和穗期对3个转Bt基因玉米品种及其相对应的受体品种进行人工接虫,分析了转Bt基因玉米对玉米螟的抗性及对产量损失的影响。结果表明：转Bt基因玉米在玉米心叶期,对一代玉米螟表现为高抗;在穗期,对二代玉米螟也表现高抗。在接虫条件下,转Bt基因玉米可以减少产量损失30%以上。因此,种植转Bt基因玉米具有明显增产作用和经济效益。     

转Bt基因抗虫玉米发展状况及对非靶标昆虫的影响

转Bt基因抗虫玉米是目前商品化进程最快的转基因作物之一,并被广泛应用于农业生产,其抗虫性和经济效益已得到普遍肯定,但外源基因的导入一定程度上改变了玉米自身的基因序列,因此转Bt基因抗虫玉米对生态安全的影响也备受关注。本文从转Bt基因抗虫玉米发展现状及对非靶标昆虫影响的角度对转Bt基因玉米的安全性研究加以综述。     

转Bt基因玉米对亚洲玉米螟的抗性及其农艺性状研究

以普通玉米杂交组合N3030、N4640为对照，通过盆栽接虫试验和田间调查，系统研究了转Bt基因玉米杂交组合N3030Bt和N4640Bt对亚洲玉米螟的抗虫效果和主要农艺性状变化。结果表明，转Bt基因玉米对亚洲玉米螟产卵无驱避作用，但花叶率、茎秆和雌雄穗的受害程度均明显轻于对照；转Bt基因玉米的生育期、株高、穗位高、穗长、穗粗等与对照差异不大，秃尖长明显短于对照，穗粒数和产量明显高于对照。     

转BcBCP1基因耐盐碱玉米对田间节肢动物群落的影响分析

利用陷阱调查法跟踪调查全生育期转BcBCP1基因耐盐碱玉米田间地上主要节肢动物群落的种类和数量。结果表明,转BcBCP1基因玉米及其受体玉米自交系黄早四主要害虫、捕食性天敌、地上跳虫的种类和数量上差异不显著,推断耐盐碱转基因玉米的种植对田间节肢动物群落短期内无显著影响。     

大豆根际未培养与培养细菌群落结构差异比较研究

根际土壤微生物群落结构是根际微生态系统中的重要组成部分,与根际养分有效性、植物生长发育及抗病性等关系密切,不同植物间、同一植物的不同基因型之间根际微生物群落结构差异较大.利用从两种基因型大豆根际土壤中直接提取和从平板培养菌落提取的微生物DNA为模板,采用细菌通用引物GC-357r和517r进行PCR扩增,对PCR产物的DGGE图谱进行聚类和主成分分析.结果表明,不同基因型大豆根际土壤未培养的细菌群落结构差异不大,而在土壤浸提液和NA培养基上形成的可培养的细菌群落结构受培养基种类和接种浓度(10-2和10-3)影响较小,但受不同大豆基因型影响而产生了差异.对DGGE条带进行分析表明,大豆根际未培养的细菌群落物种丰富度(S)和多样性指数(H)明显高于可培养细菌,说明培养过程是一个再选择的过程,在这个过程中一些微生物的信号得到放大,而大量的微生物信息缺失.对主要DGGE条带测序显示,大豆根际有三大类细菌:拟杆菌门(Bacteroidetes)、变形杆菌门(Proteobacteira)和放线菌门(Actinobacteria).变形杆菌门(Proteobacteira)中的γ-Proteobacteria、α-Proteobacteria和放线菌门(Actinobacteria),在未培养和培养细菌中都表现为优势种群.与培养细菌相比,变形杆菌门中的γ-Protebacteria和拟杆菌门(Bacteroidetes)细菌在可培养细菌中占优势,而在未培养细菌中丰度较低.结果证明,大豆根际细菌经培养后已使原有的群落结构发生改变,影响对原位细菌群落结构的认识.     

玉米株型相关基因ZmDwarf4的克隆及表达分析

以松散型玉米自交系沈137为材料,利用同源克隆法成功克隆与水稻Dwarf4基因同源的玉米Dwarf4基因。序列分析发现,该基因cDNA序列全长1 626 bp,开放阅读框1 521 bp,编码506个氨基酸,与高粱、水稻、拟南芥的氨基酸同源性分别为95%、89%和71%。Real-Time PCR分析表明,ZmDwarf4基因的表达量为3～11叶期表达量逐渐增加,9～11叶期达到最大量,12～19叶期表达量逐渐降低,推测ZmDwarf4基因正向调控玉米叶夹角的大小。ZmDwarf4基因的表达量在叶片中最高,在叶枕中最低。不同激素处理的结果表明,用植物激素（IAA）处理的样品ZmDwarf4基因的表达量始终高于对照;用Brassinolide（BR）处理的样品6叶期ZmDwarf4基因的表达量高于对照,7～10叶期该基因的表达趋势与对照基本一致,但始终低于对照;用IAA+BR处理的样品,该基因的表达趋势与对照基本一致,但高于对照。ZmDwarf4参与玉米BR生物合成途径,进而调控其相关株型建成。     

玉米ZmNRT关键基因挖掘以及氮响应基因共表达网络构建

以玉米自交系B73（V4版本）作为参考基因组,利用生物信息学方法鉴定玉米ZmNRT基因家族成员,从系统发育关系、GO（Gene Ontology）富集、基因结构和玉米不同时期及组织下的表达谱等方面全面解析玉米ZmNRT基因家族。基于qPCR实验和共表达网络分析,对玉米ZmNRT家族基因在氮响应中的作用进行系统的研究。结果表明,在玉米全基因组水平上共鉴定到 162 个 ZmNRT 基因,主要分为 ZmNRT1（62 个）和 ZmNRT2（100 个） 两大类群。GO富集发现,仅46个基因参与硝态氮转运过程,这些基因被不均等分成7个亚家族,每个家族1～15个基因。基因表达模式分析结果显示,不同生育期和组织下ZmNRT基因表达是差异的。在氮诱导下,qPCR鉴定结果显示,12个ZmNRT基因是氮响应基因,9个基因表达上调,3个基因表达下调。共表达网络结果发现,其中10个ZmNRT基因与已知的氮代谢基因存在显著共表达关系（|r|>0.8 p-value<0.05）,推测这10个基因可能是调控玉米氮代谢的关键基因。     

磷浓度处理对大豆根际土壤微生物群落结构的影响

根际土壤微生物是根际土壤生态系统的重要组成部分,与根际养分有效性、植物生长发育等关系密切.土壤磷素营养的差异可引起植物根系分泌物和根形态的变化,但对植物根际微生物的作用还少有报道.在盆栽条件下,研究了3种磷浓度处理对生长在黑土土壤上3个采样时期(v3、R1和R3期)的大豆根际微生物群落结构的影响.采用PCR-DGGE技术对细菌16S rDNA和真菌ITS片段进行解析,对DGGE图谱主成分分析(VCA).结果表明:大豆根际土壤细菌和真菌群落结构均随生育期而迁移,说明生育时期较磷浓度处理而言是影响根际微生物群落结构产生变化的主要因素;大豆根际真菌群落结构受不同磷浓度处理产生分异体现在v3期,细菌结构产生分异体现在v3和R1期,说明磷浓度处理对大豆根际微生物的影响主要表现在大豆生育的早期,随着生育时期的推进.这种差异弱化至不明显.     

我国北方潮土玉米不同生长时期古菌群落结构和丰度的变化

采用16S rRNA基因末端限制性片段长度多态性分析(T-RFLP)和实时荧光定量PCR技术,研究玉米不同生长时期土壤古菌群落组成和丰度的变化。结果表明,不同生长时期,根际土和非根际土古菌优势种群无显著变化,古菌群落组成无明显分离。乳熟期和完熟期古菌Shannon指数显著高于拔节期和抽雄期(P0.05),根际土抽雄期古菌Evenness指数显著低于其他生长时期(P0.05),非根际土不同生长时期Evenness指数差异不显著(P0.05)。土壤古菌16S rRNA基因丰度随生长时期的推进呈先升高后降低的趋势,乳熟期最高,拔节期最低,乳熟期显著高于拔节期和完熟期(P0.05),与抽雄期差异不显著(P0.05)。同一生长时期根际土和非根际土间无显著差异(P0.05)。     

玉米耐旱相关基因dbf1的序列变异分析

玉米dbf1 基因在干旱胁迫下的编码产物与DIP₁蛋白相结合,调控植物激素脱落酸（ABA）响应基因rab17 的表达,从而提高耐旱性。以B73的dbf1 基因组序列为模板,对104个玉米自交系的dbf1 基因进行测序,研究玉米dbf1 基因的序列多态性。多态性分析表明,在全长为2 118 bp的序列中共发现48个SNP和12个InDel。尽管大部分变异位点集中在非编码区,但编码区的1个InDel和3个非同义突变位点的变异可以产生氨基酸序列改变。玉米dbf1 基因的全长序列和编码区序列的变异位点可以分别将该基因划分成33和11种单倍型,并且编码7种蛋白质。在供试材料中,dbf1 基因至少经历了9次重组,中性检测表明,该基因的进化没有偏离中性选择。     

Impact of Bt-cotton on soil microbiological and biochemical attributes

Transgenic Bt-cotton produces Bt-toxins (Cry proteins) which may accumulate and persist in soil due to their binding ability on soil components. In the present study, the potential impacts of Bt- and non-Bt genotypes of cotton on soil microbial activity, substrate use efficiency, viable microbial population counts, and nutrient dynamics were studied. Two transgenic Bt-cotton genotypes (CIM-602 CIM-599) expressing cry1 Ac gene and two non-Bt cotton genotypes (CIM-573 and CIM-591) were used to evaluate their impact on biological and chemical properties of soil across the four locations in Punjab. Field trials were conducted at four locations (Central Cotton Research Institute-Multan, Naseer Pur, Kot Lal Shah, and Cotton Research Station-Bahawalpur) of different agro-ecological zones of Punjab. Rhizosphere soil samples were collected by following standard procedure from these selected locations. Results reveled that Bt-cotton had no adverse effect on microbial population (viable counts) and enzymatic activity of rhizosphere soil. Bacterial population was more in Bt-cotton rhizosphere than that of non-Bt cotton rhizosphere at all locations. Phosphatase, dehydrogenase, and oxidative metabolism of rhizosphere soil were more in Bt-cotton genotypes compared with non-Bt cotton genotypes. Cation exchange capacity, total nitrogen, extractable phosphorous, extractable potassium, active carbon, Fe and Zn contents were higher in rhizosphere of Bt-cotton genotypes compared with non-Bt cotton genotypes. It can be concluded from present study that the cultivation of Bt-cotton expressing cry1 Ac had apparently no negative effect on metabolic, microbiological activities, and nutrient dynamics of soils. Further work is needed to investigate the potential impacts of Bt-cotton on ecology of soil-dwelling insects and invertebrates before its recommendation for extensive cultivation.     

玉米秸秆深翻还田土壤细菌群落16SrDNA-PCR-DGGE分析

以秸秆深翻还田土壤为研究对象,利用嵌套式PCR和DGGE电泳技术对细菌16SrDNA V3～V5区进行扩增和产物分离,分析3个秸秆深翻处理下春玉米各生育期土壤细菌群落变化,探索高寒灌溉农区春玉米秸秆深翻还田中细菌群落结构特征。结果初步表明,秸秆还田处理细菌多样性高于常规旋耕无秸秆还田处理,这种多样性同时受玉米生育期和不同秸秆还田处理的影响。3个处理相同生育期DGGE条带数有明显的变化,玉米秸秆深翻还田两年处理（SF-II）条带数最多,其次是玉米秸秆深翻还田1年处理（SF-I）,常规旋耕无秸秆还田（CK）最少;在成熟期,SF-II条带数为12,是CK处理的两倍。玉米全生育期一直存在数种优势菌群,但各时期细菌的种类不同,整体表现为随着玉米的生长细菌种类逐渐增加,成熟期减少。秸秆深翻还田能丰富土壤微生物多样性,玉米生长中期土壤微生物多样性较丰富。     

棉铃虫对转基因抗虫玉米的行为反应

棉铃虫Helicoverpa armigera(Hübner)是Bt玉米的重要靶标害虫之一。规避行为可以减少害虫与Bt玉米的接触而降低生理抗性选择压力。通过比较棉铃虫对Bt玉米与常规玉米的行为反应评估棉铃虫对Bt玉米的行为规避能力,产卵选择结果显示,棉铃虫成虫在Bt玉米上的落卵量显著低于常规玉米;无选择条件下,仍然出现Bt玉米上的落卵量显著低于常规玉米的现象。幼虫实验结果显示,棉铃虫初孵幼虫在Bt玉米植株上的居留时间显著短于在常规玉米上的居留时间。研究结果证明,棉铃虫对Bt玉米具有一定的行为规避能力。     

Influence of transgenic hybrid rice expressing a fused gene derived from cry1Ab and cry1Ac on primary insect pests and rice yield

Although dozens of transgenic Bacillus thuringiensis (Bt) rice lines have been developed, none of them has been released to farmers. Under field conditions, we evaluated the influence of a hybrid Bt rice on the primary rice insect pests and rice yield in 2005 and 2006. Four treatments were evaluated, including Bt and non-Bt rice treated with insecticides when necessary, and unprotected Bt and non-Bt rice. Unprotected Bt rice exhibited stable and high control of the three primary lepidopteran pests, Chilo suppressalis Walker, Tryporyza incertulas Walker and Cnaphalocrocis edinalis Güenée. Under unprotected conditions, larval densities of these three pests in Bt plots decreased by 87.5–100% compared to those in non-Bt plots, and percentages of damaged stems and leaves remained less than 0.6% during the entire rice growing season. In early rice growth stages, populations of two important planthoppers, Nilaparvata lugens Stål and Sogatella furcifera Hovarth, were significantly affected only by protection level (protected vs unprotected). However, in late rice growth stages (filling and maturing), densities of planthoppers were significantly affected both by protection level and by rice type (Bt vs non-Bt), and densities of N. lugens were significantly higher in Bt plots than in non-Bt plots under unprotected conditions. Pesticide sprays were reduced by 60 and 50% in protected Bt vs protected non-Bt plots in 2005 and 2006, respectively. Yield of unprotected Bt rice increased by 60–65% compared to unprotected non-Bt rice, but decreased by 28–36% compared to protected Bt rice. These results show that Bt rice increased yield greatly, but still required pesticide sprays to avoid losses caused by non-target insect pests.     

Transgenic Bt corn varietal resistance against the Mexican rice borer,Eoreuma loftini (Dyar) (Lepidoptera: Crambidae) and implications to sugarcane

The Mexican rice borer, Eoreuma loftini (Dyar), attacks crops including corn, Zea mays L., rice, Oryza sativa L., sorghum, Sorghum bicolor (L.) Moench, and sugarcane, Saccharum spp. Strongly resistant varieties of any kind, native or otherwise, have not been identified. A field plot corn variety test using two transgenic Bt varieties, Pioneer 31G71, expressing the Cry1F insecticidal protein, and Golden Acres 28V81, expressing the Cry1A.105, Cry2Ab2, and Cry3Bb1 insecticidal proteins, and two non-Bt controls, Dekalb DKC 69-72 and BH Genetics 9050, all four commonly grown in the Lower Rio Grande Valley of Texas, showed that, although oviposition preference was not affected, 28V81 resisted larval stalk boring to the extent that Mexican rice borer injury was almost non-existent. Pioneer 31G71 was infested nearly as much as the controls, but larval development to adulthood was reduced by ≈70%. Rearing larvae on 5, 50, 500, and 5000 μg of corn leaf tissue per ml of artificial diet showed that, while the three lowest concentrations did not affect larval growth and development, the high concentration of 28V81 reduced survivorship to the pupal stage, decreased weight of 4-wk-old larvae, and prolonged development to pupation. Lower numbers of pheromone trap-captured adults at the edges of commercial Bt and non-Bt corn fields showed that populations were lower at the Bt cornfields, suggesting a lesser rate of adult production. Because corn is a preferred host plant over sugarcane, sorghum or rice, use of resistant transgenic Bt corn varieties will likely protect the crop from the substantial injury that can be caused by the pest. This study also suggests that Bt genes might result in similarly strong resistance when inserted in other vulnerable crops such as sugarcane.     

Theoretical and empirical assessment of a seed mix refuge in corn for southwestern corn borer

Larval movement in seed mixes of Bt and non-Bt corn (Zea mays L.) can potentially increase the rate at which resistance evolves compared to a structured refuge. Pyramided Bt crops with multiple efficacious Bt toxins make it less likely that interplant larval movement will be successful within a seed mix field but high levels of larval movement may decrease refuge value as larvae moving from non-Bt plants die on neighboring Bt plants. Estimates of refuge productivity obtained from multi-site field studies evaluating larval movement and survival of the southwestern corn borer (SWCB), Diatraea grandiosella Dyar, in MON 89034 seed mix refuge fields were compared to predictions from model simulations of larval movement within a seed mix refuge. Results from field studies and modeling simulations of interplant movement show that seed mix refuge was a productive source of susceptible SWCB and that successful interplant movement by larvae within a seed mix refuge was unlikely to occur at frequency that would increase the fitness of resistant heterozygotes or low level resistance mechanisms. Combined, these results indicate that MON 89034 with a 5% seed mix refuge is a durable IRM tactic for SWCB across a range of larval movement and refuge assumptions and show that a 5% seed mix refuge can delay resistance longer than a similar-sized structured refuge given the nature of grower compliance with refuge requirements.     

中国北方玉蜀黍黑粉菌交配型a位点鉴别及遗传多样性分析

利用特异性引物对2017年采自7个省(自治区)的84份玉米瘤黑粉病菌进行交配型a位点鉴别,采用相关序列扩增多态性(ISSR)分子标记技术对供试菌株进行遗传多样性分析,分析我国北方玉米主要产区玉蜀黍黑粉菌交配型a位点基因型种类及其遗传多样性水平。结果表明,玉蜀黍黑粉菌交配型a位点基因型主要有mfa1、mfa2和mfa1+mfa2三个种类,84株菌株中,mfa1基因型和mfa1+mfa2基因型均有31株,占鉴定株数的36.9%;22株为mfa2位点,占鉴定株数的26.2%。通过筛选出的8条ISSR引物共扩增出61条清晰条带,特异性条带55条,多态性位点所占比例为90.2%。UPGMA聚类分析结果表明,供试菌株遗传相似系数介于0.59～0.92,在遗传相似系数为0.65时供试菌株被分为6个类群,表现出丰富的遗传多样性。ISSR揭示的玉蜀黍黑粉菌遗传多样性、交配型a位点基因型以及地理来源之间的关系分析结果表明,菌株的遗传类群和交配型a位点基因型之间无密切关系,并且两者与菌株地理来源也无明显相关性。     

我国北部及中东部地区玉米根际土壤中寄生线虫种类调查研究

对我国北部及中东部9省(直辖市)的57市(县)采集玉米根际土壤样品,从92份土样中共分离到17属植物寄生线虫,即垫刃属(Tylenchus)、丝尾垫刃属(Filenchus)、杆垫刃属(Rhabdotylenchus)、平滑垫刃属(Psilenchus)、叉针属(Boleodorus)、巴兹尔属(Basiria)、矮化属(Tylenchorhynchus)、短体属(Pratylenchus)、小环属(Criconemella)、茎属(Ditylenchus)、纽带属(Hoplolaimus)、盾属(Scutellonema)、盘旋属(Rotylenchus)、螺旋属(Helicotylenchus)、拟盘旋属(Pararotylenchus)、真滑刃属(Aphelenchus)、滑刃属(Aphelenchoides)。其中,矮化属线虫分布较广,占样本总数的58.70%;螺旋属线虫的相对丰度最大,占线虫总数的47.53%,这两属线虫为玉米根际土壤中的优势属。对不同地区线虫种群进行分析得出,螺旋属线虫为山东、安徽、江苏及河南省的优势属,矮化属线虫为黑龙江、吉林、辽宁、北京和河北省(市)的优势属,垫刃属线虫是吉林省的优势属,短体属线虫为辽宁和河南两省的优势属。     

转基因小麦根际土壤中荧光假单胞菌数量的变化

荧光假单胞菌(Psdeuomnoda fluoerncnet)是一种土壤中十分常见的细菌,广泛应用于生物防治,其数量变化能较准确地反映土壤环境的变化,因此在转基因小麦环境安全评价研究中可以将荧光假单胞菌的数量变化作为检测转基因小麦种植对整个根际土壤环境影响的指标之一。本研究选取荧光假单胞菌的蛋白编码基因gyrB作为靶标基因,设计特异性较好的引物,利用该引物建立了基于实时荧光定量PCR(RFQ-PCR)方法的转基因小麦根际土壤荧光假单胞菌数量的检测技术体系。结果显示,定量标准曲线相关系数达0.99以上,且呈现单一熔解曲线峰值。应用该体系对种植于河南新乡及江苏六合的转TaDREB4基因抗旱小麦(简称TB4)及其受体小麦济麦19根际土壤荧光假单胞菌进行检测,结果表明,各时期转基因小麦和非转基因小麦各生育期根际土壤中荧光假单胞菌的拷贝数无显著差异。说明TB4种植对根际土壤中的荧光假单胞菌数量无显著影响。     

Evaluation of European corn borer Ostrinia nubilalis (Lepidoptera: Crambidae) larval movement and survival in structured and seed blend refuge plantings

Blending Bacillus thuringiensis (Bt) corn and non-Bt corn seeds in the same bag is a convenient and easier compliance solution to satisfy refuge regulations. However, there has been considerable debate and concern about larval movement of the target insects in a seed blend because of its potential effect on the development of resistance. Reported here are studies to determine the larval movement of European corn borers, Ostrinia nubilalis (Hübner), in a seed blend compared to a structured refuge and their effects on survival, feeding injury and larval fitness. Also evaluated is the relative performance of a purple-seeded corn hybrid as a surrogate host plant for tracking gene flow. Seed blend and structured refuge arrangements of Agrisure GT/CB/LL Bt corn expressing Cry1Ab and isoline plants were evaluated at two locations in 2013 and 2014 by manually infesting refuge plants with first and second generation corn borer larvae. The majority of leaf and stalk injury was recorded on the refuge plant, which amounted to 66–83% of the total tunnel length. In both refuge arrangements, plant injury and the number of larvae recovered significantly decreased on neighboring plants with increasing distance away from the refuge plant in the same row. The relative decline in injury was much more pronounced in the seed blend, with neighbor Bt plants experiencing very minor tunneling or no injury at all due to the high dose trait. Eighty-nine percent fewer live larvae were also found on neighbor Bt plants in the seed blend compared to those found in the structured refuge plots. Larvae that moved to Bt plants as well as older instars that were manually placed on Bt plants failed to complete development. Comparison of the number of tunnels and larvae recovered per refuge plant indicated that the seed blend may not produce as many susceptible individuals as those produced in a structured refuge arrangement. Although the purple-seeded hybrid showed lower levels of ear and stalk injury, it could serve as an appropriate surrogate refuge plant to track gene flow.