双价抗虫节水抗旱稻新材料旱恢3CA的创制

绿色超级稻的育种目标是少打农药、少施化肥、节水抗旱和优质高产。为培育“少打农药,节水抗旱”的资源节约型、环境友好型水稻品种,本研究以T1C-19、T2A-1作为cry1C^*和cry2A的基因供体,与节水抗旱稻恢复系旱恢3号品种杂交,利用分子标记辅助选择技术对目标基因进行筛选,最终获得同时含有cry1C^*和cry2A基因的稳定株系。通过PCR以及对Cry1C^*和Cry2A蛋白的定性和定量检测,结合正常管理田间农艺性状考察及不防治螟虫田间管理的抗虫性鉴定,筛选出双价抗虫节水抗旱稻新恢复系旱恢3CA。结果显示,对照旱恢3号与旱恢3CA在农艺性状上无显著差异,但早恢3CA高抗稻纵卷叶螟。本研究结果为培育既节水抗旱又抗虫的水稻新品种奠定了材料基础。     

Cry1Ab转基因水稻的杂种优势表现及抗虫性鉴定

以携带Cry1Ab基因的"克螟稻"为抗虫供体亲本,将目标基因转导到优良恢复系"R6547"、"R818"中,配制的杂交稻组合表现良好的杂种优势水平,克服了"克螟稻"在常规粳稻育种实践中常出现的农艺性状差、前期生长势弱的缺陷。可溶性蛋白含量检测结果显示目标基因在杂交稻中仍能高水平的表达,并在人工接虫和自然发生条件下对稻纵卷叶螟、二化螟等鳞翅目害虫表现优良抗性。     

Cry1Ac蛋白降解菌株FJSB3的分离鉴定及降解特性

从转Cry1Ac基因水稻种植田土壤中,分离纯化得到1株能高效降解Cry1Ac蛋白的细菌FJSB3。通过表型特征、16S rDNA扩增和电镜观察,初步鉴定FJSB3为寡养单胞菌（Stenotrophomonas sp.）。FJSB3发酵液能降解粗Bt蛋白。通过单因素试验确定FJSB3降解水稻秸秆中Cry1Ac蛋白的最适条...     

泥鳅生长及抗氧化 解毒酶系统对水体中转Cry1Ab/Ac基因水稻残遗物的响应

在泥鳅养殖水体中添加稻秆粉模拟水稻残遗物生境,研究了泥鳅生长和肝胰脏抗氧化酶(SOD、CAT)与解毒酶(GST)活性对转Cry1Ab/Ac基因水稻‘华恢1号’(HH1)的响应。设计以HH1稻秆粉10 mg·L?1、50 mg·L?1、100 mg·L?1和200 mg·L?1 4个梯度浓度处理泥鳅为试验组,以非转Bt基因水稻‘明恢63’(MH63)稻秆粉处理组为阴性对照,不加稻秆粉的基础饲养组为空白对照。结果显示:在4种稻秆粉浓度下,HH1组与MH63对照组泥鳅的特定生长率、肥满度、内脏系数及SOD、CAT和GST酶活性均无显著差异(P0.05);与空白对照比较,稻秆粉浓度升高对泥鳅生长的抑制逐渐增强,当浓度达到200 mg·L?1时,HH1组和MH63对照组泥鳅的特定生长率、内脏系数与CAT活性降低。研究结果表明,水体中低含量的转融合基因Cry1Ab/Ac水稻HH1稻秆粉对泥鳅的生长与生理酶活性没有明显影响,高浓度HH1和MH63稻秆粉均使泥鳅的生长和生理酶活性显著降低,这可能与养殖水体中浓度较高的悬浮稻秆粉妨碍了泥鳅的呼吸和滤食,及稻秆粉的分解降低了水体p H和溶氧量有关。     

转基因高粱Cry1Ab蛋白含量的比较研究

通过农杆菌介导法将杀虫晶体蛋白基因cry1Ab导入高粱恢复系115中,共获得13个独立的转基因株系,26株转基因植株,转化率为5.1%;GUS活性、PCR、Southern和Western杂交分析表明,此基因已整合进高粱基因组并得到正确表达。利用ELISA试剂盒测定Cry1Ab蛋白含量,结果表明,不同转基因植株的Cry1Ab蛋白含量有明显差异,最高可达0.850 μg/g,占可溶性蛋白的0.016%,还有一些转基因植株中不能表达;同一转基因植株的不同组织中表达量有明显差异,其顺序为：叶>颖壳>籽粒>根>茎;不同部位的叶片也有差异,其顺序为：中部叶>基部叶>新叶。     

不同发育时期转基因高粱不同株系Cry1Ab蛋白含量的比较

高粱的害虫种类众多,影响着产量与品质,培育抗虫品种是重要的育种目标,但目前高粱中缺乏有效的抗螟虫资源,难以通过常规育种培育出抗螟虫的品种。张明洲(2002)成功地将来自Bt密码子优化的抗虫基因cry1Ab转入高粱中,并经农业部批准进入中间试验阶段。为有效和合理地利用这一抗虫     

水稻恢复系辐恢838及其衍生系的选育和应用

采用辐射诱变、杂交及逆境温度筛选相结合的综合育种方法,成功育成了辐恢838及其衍生恢复系辐恢718、辐恢305、中恢218、绵恢3728、糯恢1号等12个恢复力强、配合力高、抗逆性好的恢复系。利用这些恢复系与野败、冈型、D型、印水型、夜公型等胞质不育系组配育成的43个组合,先后通过了国家或省农作物品种审定。这些品种在大面积种植过程中表现出结实率高、较耐低温冷害和高温热害、适应性广、高产稳产的特点,累计种植面积4000万hm2 以上。其中II优838是继汕优63以后我国应用时间长、种植面积较大的著名品种。2005年成为国家籼稻区域试验对照组合,也是我国近年来主要的出口越南东南亚等国家的杂交水稻品种。     

不同光质LED光源对转基因杨树叶片Bt毒蛋白含量及叶绿素荧光参数的影响

为探明不同光质对杨树幼苗生长的影响,以转基因741杨株系Pb29为试材,利用LED精量调制光源,设置红光、蓝光、红蓝光、白光4个处理,测定不同光质处理下Pb29的Cry1Ac毒蛋白含量、形态指标、光合色素含量及叶绿素荧光参数。结果表明,蓝光处理下的Cry1Ac毒蛋白的表达量最大,白光次之。除了叶片长/宽的值在蓝光处理下最大外,红蓝光处理下的741杨叶片数、叶片长、叶片宽、叶面积及株高均高于其它处理。除叶绿素b的含量在红光处理下最大外,叶绿素a、叶绿素a+b、类胡萝卜素含量及叶绿素a/b比值均在白光处理下最大,红蓝光次之,且白光与红蓝光间无显著差异。叶绿素荧光参数(ΦPSⅡ、ETR、q P)在蓝光和红蓝光处理下高于其它处理,在白光和红蓝光处理下的Fv/Fm高于其它处理,q N在红光处理下最大。由此可知,不同光质对杨树幼苗的生长具有调控作用。本试验结果为深入研究不同光质对杨树的生长发育进行调控提供了科学依据。     

转Cry1Ab/Ac基因水稻对稻田底栖动物群落的影响

转Bt基因抗虫水稻的成功研发,为有效控制鳞翅目害虫对水稻的危害提供了一条最为有效、经济的途径。稻田底栖动物群落是转Bt基因抗虫水稻的重要非靶标生物。为了解转Bt基因水稻对稻田底栖动物群落的安全性,以转Cry1Ab/Ac基因抗虫水稻‘华恢1号’(简写为HH1)为材料,以其非转基因亲本‘明恢63’(简写为MH63)为对照,以稻田底栖动物群落为指示生物,运用群落学方法,通过连续2年在江西南昌的大田试验,研究了转Cry1Ab/Ac基因抗虫水稻对稻田底栖动物群落的影响。结果显示:2012年在HH1与MH63的稻田中分别采集到底栖动物22种与25种,其中两生境中相同物种19种,两群落物种组成相似度为0.808 5,有7种优势种,优势种相似度为0.833 3。2013年在HH1与MH63稻田中分别采集到底栖动物26种与28种,两类稻田相同底栖动物22种,两群落物种组成相似度为0.814 8,有6种优势种,优势种相似度为1.000 0。2012年与2013年群落总体分析与时间动态分析均显示,两类稻田中底栖动物的物种丰富度、个体数量、多样性指数、均匀性指数及优势集中性指数变化趋势相似,且均无显著差异。结果表明:连续种植2年转Cry1Ab/Ac基因水稻HH1对南昌稻田底栖动物群落无明显的负作用。     

南非冬闲期表施和混施入Bt层玉米落叶和Cry1Ab蛋白的分解研究

Unintended effects of genetic modification on chemical composition of Bt maize leaf litter may have impacts on its decomposition. In most agricultural systems in South Africa, maize litter is either left on the soil surface or incorporated into the soil during tillage. A litterbag experiment, using leaf litter of three maize hybrids （DKC80-12B, DKC80-10 and DKC6-125）, was carried out at the University of Fort Hare Research Farm, South Africa, to determine the effects of genetic modification on decomposition of maize leaf litter when left on the soil surface under field conditions between July and November, the normal fallow period, in 2008. Another litterbag experiment was conducted at the University of Fort Hare Research Farm and Zanyokwe Irrigation Scheme, South Africa, using leaf ~itter of two maize hybrids genetically modified with the erylAb gene （MONS10）, DKC75-15B and PAN6Q-3OSB, and their corresponding near isolines, CRN3505 and PAN6Q-121. The degradation of CrylAb protein in the litter, both surface-applied and soil-incorporated, was also investigated. Decomposition of Bt maize litter was similar to that of non-Bt maize litter both when applied on the surface and when incorporated into soil. Soil-incorporated litter, as well as its CrylAb protein, decomposed faster than that applied on the surface. The leaf litter C：N ratios of PAN6Q-308B and PAN6Q-121 were similar throughout the study, whereas those of DKC75-15B and CRN3505 declined by similar amounts during a 12-week period. These findings suggested that decomposition of leaf litter of Bt maize, with the MON810 event, was not affected by maize genetic modification, and that the CrylAb protein broke down together with plant leaf litter during the winter fallow regardless of whether the litter was applied on the soil surface or incorporated into soil.     

Effects of temperature, water content and pH on degradation of Cry1Ab protein released from Bt corn straw in soil

We investigated the effects of soil temperature (15 °C, 25 °C, 35 °C), water content (20%, 33%, 50%) and pH (4.5, 7.0, 9.0) on the degradation of Cry1Ab protein released from the straw of Bt corn varieties 34B24 and 1246 × 1482 both expressing Cry1Ab protein. Our results showed that Cry1Ab protein released from both 34B24 and 1246 × 1482 straw was degraded in a similar way in all treatments, which demonstrated a rapid decline in the early stage but a slow decline in the middle and late stages. In the late stage (180 days after the experiment commenced) 0.03%-1.51% and 0.02%-0.91% of initial Cry1Ab protein released from 34B24 and 1246 × 1482 straw was detected in soil. In addition, degradation dynamics of Cry1Ab protein under different environmental conditions was well described by the shift-log model. DT₅₀ of Cry1Ab protein released from 34B24 and 1246 × 1482 straw was 0.97-9.97 d and 0.75-10.89 d, respectively, and DT₉₀ was 4.66-162.45 d and 6.44-57.46 d, respectively. The results suggested that soil temperature had significant effects on the degradation of Cry1Ab protein, with a higher degradation rate at higher temperature, but soil water content and pH had no obvious effects on the degradation of Cry1Ab protein.     

苏云金芽胞杆菌营养期杀虫蛋白vip3A-LS1基因的克隆与表达

从土壤中分离得到营养期对鳞翅目害虫高毒的苏云金芽胞杆菌(Bacillus thuringiensis,Bt)LS1菌株,分子检测该菌株中存在营养期杀虫蛋白基因,进行了该基因的克隆和表达。设计全长基因引物扩增得到了约2.3kb的靶片段,序列分析证实为新vip3A基因,命名为vip3A-LS1,GenBank登录号为DQ016968,Bt基因命名委员会命名为Vip3Aa22。该基因推导蛋白与其它已知同源蛋白有8个氨基酸不同。构建vip3A-LS1基因的表达载体,SDS-PAGE检测约88kD目的蛋白大量表达;生物测定表明,胞内可溶性蛋白对棉铃虫(Helicoverpa armigera)和甜菜夜蛾(Spodoptera exigua)活性最高。纯化蛋白对初孵棉铃虫和甜菜夜蛾的LC50分别为73.6和32.2μg/g。     

Effects of physical and chemical properties of soils on adsorption of the insecticidal protein (Cry1Ab) from Bacillus thuringiensis at Cry1Ab protein concentrations relevant for experimental field sites

The adsorption of the insecticidal Cry1Ab protein of Bacillus thuringiensis (Bt) on Na-montmorillonite (M-Na) and soil clay fractions was studied. The aim of this study was not to find the adsorption capacity of the soils from the experimental field site, where Bt corn (MON810) was cultivated, but rather to characterize the adsorption behavior of the Cry1Ab protein at concentrations typically found at experimental field sites. In kinetic experiments, the Cry1Ab protein adsorbed rapidly (<60 min) on M-Na. As the concentration of M-Na was varied and the added Cry1Ab protein concentration was kept constant (20 and 45 ng ml⁻¹), the adsorption per unit weight of Cry1Ab protein decreased with increasing concentrations of M-Na. Adsorption of Cry1Ab protein on M-Na decreased as the pH value of the suspension increased. All adsorption isotherms could be described mathematically by a linear regression with the parameter k, the distribution coefficient, being the slope of the regression line. Although their mineralogical composition was nearly identical, the soil clay fractions showed different k values. The different k values were correlated with the physical and chemical properties of the soil clay fractions, such as the organic carbon content, the specific external surface area, and the electrokinetic charge of the external surfaces of the clays, as well as with the external surface charge density. An increase in the amount of soil organic matter, as well as an increase in the electrokinetic external surface charge of the soil clays, decreased the distribution coefficient k. An increase of the specific external surface areas of the soil clays resulted in a higher distribution coefficient k.Less than 10% of adsorbed Cry1Ab protein was reversibly adsorbed on the soil clays and, thus, desorbed. The desorption efficiency of distilled water was higher than that of a solution of CaCl₂ (2.25 mmol) and of dissolved organic carbon (50 mg C l⁻¹).     

快长、少畸形香鱼新品种“浙闽1号”的选育

香鱼(Plecoglossus altivelis)“浙闽1号”是以2002年采捕自浙江宁海凫溪的野生香鱼为基础群体,采用群体选育技术,以生长速度和体形特征为选育指标,经7代连续选育而获得的水产新品种.该品种在相同养殖条件下,9月龄时体长生长要比当地商品苗快11.36％以上,体重生长快25.92％.2012～2014年在浙江宁海和福建南靖两处进行中试,工厂化养殖面积达4.5万m2,9月龄鱼存活率比当地商品苗提高30％以上,畸形率接近于零,平均增收38.98％.本文介绍了“浙闽1号”选育过程、品种特征和中试情况,为该品种的进一步推广养殖以及其他水产新品种的选育提供信息和资料借鉴.     

城市湖泊富营养化及氮磷平衡研究——以湖北省黄石市青山湖1#湖区为例

[目的]监测分析湖北省黄石市青山湖上游区域水质状况,为其生态修复提供依据。[方法]以青山湖1~#子湖区为例,采用综合营养状态指数法对其富营养化水平进行评价,依据物质平衡原理估算出青山湖上游区域氮磷收支状况。[结果]①青山湖上游区域水体总氮、总磷年均浓度为2.45,0.200 mg/L;②青山湖上游区域水体达到中度富营养化水平;③上游来水是氮磷入湖主要来源;④氮入湖总量6 115.40 kg/a,出湖总量1 939.51 kg/a;⑤磷入湖总量411.14 kg/a,出湖量总量198.21 kg/a;⑥总磷、总氮截留率分别为68.28%,51.79%。[结论]为防止青山湖上游区域富营养化程度加重,建议对上游来水采用生态浮床的措施,在净化水质,达到外源清水入湖的同时还能改善景观。在排口附近湖水区域,可采取高效原位强化净化处理措施。