利用花粉管通道技术导入大豆抗病虫目的基因

采用花粉管通道技术,将几丁质酶基因导入14个大豆品种,共导入1125朵花,花朵的成活率为47.8%,将Bt基因导入11个大豆品种,共导入260朵花,花朵的成活率为40.8%。8个转Bt基因品种中的D1代共收获192粒种子,获得幼苗132株,出苗率为68.75%,把转Bt基因的132株植株用X-Gluc溶液检测,没有发现阳性反应,将转Bt基因的132株植株进行PCR检测,得到5株阳性转化植株,对D1代获得的5株阳性转化植株的D2代,进行PCR检测,得到D2代稳定遗传的阳性转化植株2株。     

发根农杆菌介导几丁质酶基因和β-1,3-葡聚糖酶基因转化烟草的研究

用冻融法和三亲交配法将具有几丁质酶基因、β—1,3—葡聚糖酶基因和卡那霉素抗性标记基因的质牲pBLGC导入具有Ri质粒的发根农杆菌中。用叶盘法转化烟草的三个品种,经Kan抗性筛选获得126株再生植株,对119株再生植株分别以启动子CaMV35S、几丁质酶基因和β—1,3—葡聚糖酶基因的引物进行PCR检测,其中几丁质酶基因至阳性的植株有72株,β—1,3—葡聚糖酶基因至阳性的有47株,具有几丁质酶基因和β—1,3—葡聚糖酶基因的植株有36株。对转化的24株转基因经PCR-Southern杂交,结果均至阳性。实验结果表明,外源基因的转化频率与植物品种、外源基因片段的大小有关,几丁质酶基因和β—1,3—葡聚糖酶基因分别或共同整合到植物基因组中。     

几丁质酶基因导入西瓜植株及其抗病性鉴定研究

利用西瓜有性生殖过程 ,将携带有外源几丁质酶基因的质粒DNA涂抹在授粉后的柱头上 ,使其沿花粉管通道进入到生殖细胞 ,得到转化处理种子。转化T1代植株经除草剂Basta筛选和PCR扩增获得转化植株。对T3 代株系进行田间自然发病和人工接种鉴定获得 3个抗枯萎病的株系。结果表明 :几丁质酶对镰刀菌引起的西瓜枯萎病有一定的抑制作用 ,利用花粉管通道法直接导入西瓜活体植株的技术是可行的。     

转β-1,3-葡聚糖酶基因和几丁质酶基因棉花

为了提高棉花的抗枯、黄萎病的能力,用花粉管通道法,将烟草β-1,3-葡聚糖酶基因和菜豆几丁质酶基因导入棉花,获得了抗卡那霉素的转化植株。经PCR、PCR－Southern Blot检测,证明两种基因已插入到棉花基因组中。     

将抗真菌病基因导入马铃薯的研究

实验采用微束激光穿刺技术和农杆菌介导方法,将携带有Ⅰ型烟草β－１,３－葡聚糖酶基因和Ⅰ型菜豆几丁质酶基因的ｐＢＬＧＣ质粒导入马铃薯（Ｓｏｌａｎｕｍ ｔｕｂｅｒｏｓｕｍ Ｌ．）品种“津引８号”。微束激光穿刺技术转化中,获得了７株卡那霉素抗性再生苗,分子检测证明,其中有１株为菜豆几丁质酶基因单基因转化植株。在农杆菌介导方法中得到了１３株卡那霉素抗性再生苗,其中有３株为烟草β－１,３－葡聚糖酶基因单基因     

几丁质酶基因表达载体构建及转化烟草的研究

构建了一个含有多个调控序列的带有几丁质酶基因的植物表达载体,通过发根农杆菌的Ri质粒介导转化烟草的三个品种：K326,RG11,VA116,在卡那霉素抗性培养基上筛选,获得了7株再生植株,以PCR检测、DNA斑点杂交证明表达载体构建成功,几丁质酶基因导入烟草并整合到烟草基因组中。     

IPsaur-ipt基因转化番茄的过氧化物酶同工酶分析

将整合在Ｔｉ质粒上的Ｐｓａｕｒ基因（生长素调节基因和细胞激动素合成酶基因的融合基因）通过花粉管通道法导入番茄品种中蔬四号、强力米寿、粉红甜肉、ＵＳ—Ⅰ、ＵＳ—Ⅱ中。对Ｄ１代转基因番茄用聚丙稀酰凝胶电泳方法进行过氧化物酶同工分析。结果表明,转基因番茄与对照植株的过氧化物酶同工酶谱带有明显差异,谱带多少和强弱都有所不同,而且相应地出现了植株形态学上的差异。间接验证了外源ＤＮＡ的导入。说明同工酶分析方法和田间形态学检测可作为转基因植株早期筛选的有效方法     

马铃薯X病毒外壳蛋白基因在转基因烟草植株中的表达及抗病

通过根癌农杆菌介导的叶盘法,将马铃薯Ｘ病毒外壳蛋白基因导入烟草细胞并获得大量再生的转基因烟草植株。经胭脂碱检测、酶联免疫分析和Ｗｅｓｔｅｒｎ ｂｌｏｔ分析,证明已获得了具有马铃薯Ｘ病毒外壳蛋白基因表达的转基因烟草。用２μｇ／ｍｌ的ＰＶＸ磨擦接种烟草,８天后进行观察,发现对照植株开始发病,而转基因烟草植株在２０后才开始发病,有的转基因烟草植株在整个生长期中都不得病。对ＰＶＸＣＰ基因表达量较高的４株植     

几丁质酶—葡聚糖酶双介基因导入滇型杂交稻恢复系提高稻瘟病抗性的研究

将含有菜豆（Phaseolus limensis)几丁质酶基因和烟草（Nicotiana tabacum)β-1,3-葡聚糖酶基因的pBLGC(16.5kb)质粒用基因枪法导入滇型杂交稻（Oryza sativa L.ssp.japonica)恢复系“南29”中,总计获得93个转化再生植株,以β-1,3-葡聚糖酶基因制备探讨对T1代株系进行Southern杂交分析,17个株系为杂交阳性,证实外源基因完整结构已整合到水稻基因组中;连续多代的PCR追踪检测证实外源基因已遗传至T4代;对所获得的6个PCR检测阳性T4代品系进行了稻瘟病菌（Magnaporthe grisea)生理小种接种鉴定和稻瘟病大田诱发鉴定,结果表明,转基因品系对稻瘟病的抗性较受体对照大幅度提高,获得了稻瘟病新抗源,但不同品系稻瘟病抗性并不相同。     

导入bar基因的水稻新植株可遗传

江苏省农业科学院粮作所王才林、赵凌、宗寿余等和中科院上海生化所龚蓁蓁等9位科研人员,用花粉管通道法将bar基因导入水稻获得可遗传的转基因植株。亦即他们利用花粉管通道法将抗Basta除草剂的bar基因导入水稻品系E32,获得转基因植株。但在T0代仅表现为部分抗性,T1代全部获得了抗性,T3代全部转基因植株能充分表达对Basta除草剂的抗性。通过对转基因植株后代进行PCR分析,证实了bar基因已整合到受体植株的基因组之中,以后又经过遗传分析表明,bar基因能在有性生殖过程中传递给后代植株,并在T3代开始就可分离出抗性一致的稳定株系。目前,其…     

低能离子束介导外源基因转化烟草的研究

以烟草ＮＣ－８９种子为材料,用显微扫描电镜（ＥＳＭ）和电子自旋共振（ＥＳＲ）波谱仪研究氮离子束对烟草种子表面的刻蚀作用及能量沉积产生自由基的间接效应,为离子束介导转移外源基因提供了形态结构依据。将烟草种子用２０Ｋｅｖ的氮离子束处理后,浸入含有ＰＢⅠ１２１质粒的缓冲介质中,在含有卡那霉素１００ｍｇ／Ｌ的ＭＳ０培养基上继代筛选,得到３株抗性植株。取抗性植株的叶片,经组织培养后得到再生抗性植株。经过ＰＣＲ及ｓｏｕｔｈｅｒｎ杂交分析,证明外源基因已转入烟草。     

烟叶主要生物酶活性变化规律及其在烟叶发酵中的应用

烟叶中的生物酶在烟叶的生长及发酵过程中起着重要作用.他们能够促进烟叶内部有机物质的分解与转化,调整烟叶中各种化学成分的比例,提高烟叶中香气成分的形成和积累,从而提高烟叶的综合品质.综述了烟叶生长、加工过程中淀粉酶、蛋白酶、多酚氧化酶的活性变化规律,以及外加生物酶在烟叶发酵中的应用现状.     

Some virus and virus-like diseases of tobacco,tomato, papaya,and rubber tree in vietnam and cambodia

In Vietnam a green strain of tobacco mosaic virus was isolated having TIP 89°C (10 min) and causing systemic necrosis in tobacoo ‘Xanthi-nc’ and sometimes also inDatura stramonium. In symptomless tomato plants an elongated virus belonging apparently to the Carlavirus group (NL 630 nm) was found. In papaya trees showing severe symptoms of mosaic and/or ringspot elongated virus particles (NL 730 nm) were observed; this virus being apparently a member of the Potyvirus group, resembled as far as its symptoms in papaya are concerned, the papaya ringspot or the distortion ringspot. In Cambodia some young rubber trees showed malformed leaves (esp. edges and veins) with yellow discolorations along the veins. Such leaves contained elongated virus-like particles (rigid or slightly flexible) of various length (60 to 880 nm), so that their normal length (NL) could not be established precisely. Particles 120 to 150 nm long occurred very frequently.     

Dependence of the Effectiveness of Induced Resistance on Environmental Conditions

Resistance induced by microbial metabolites against powdery mildew on barley always proved to be highly effective particularly under practical conditions in field. The results obtained in the present study demonstrate that the successful induction of resistance requires a conditioning of plants by the environmental conditions acting prior to the application of the inducer. If plants were grown in the greenhouse or outdoors, the infection densities were efficiently reduced after the treatment with the inducer. In contrast to this the disease intensities were not reduced on plants grown at constant temperature, light and humidity conditions. Frequently changing temperatures and the daily courses of temperature, normally occurring in greenhouse or outdoors, were substantiated as the most important factors for a successful induction of resistance. The conditioning of the plants under these temperature conditions depended on light energy, especially in just developing leaves. The dependence of the induced resistance on the environmental conditions was proved with other host-pathogen-relationships as well as with barley and powdery mildew.     

The uptake,transport and metabolism of the bioregulator 2-(3,4-dichlorophenoxy) ethyldimethylamine in guayule

The bioregulator 2-(3,4-dichlorophenoxy) ethyldimethylamine was applied to five-month-old summer and winter guayule plants. Uptake of this molecule depended on the presence of viable trichomes and a well-developed cuticle, in the leaves. Winter plants absorbed the bioregulator more successfully than summer plants. The stem proved to be an active absorption site in young plants. Six days after bioregulator application, transport of the molecule was restricted to the lower stem in summer plants, and stem and leaves in winter plants. Transport was governed by the availability and development of conduits. The intact molecule was recovered two days after application but was not detectable after 4 and 6 days indicating that it is metabolized fairly rapidly. The significance of these findings is discussed in terms of the use of bioregulators to stimulate rubber production in guayule plants.     

转果聚糖蔗糖转移酶基因( Sac B)美丽胡枝子的获得

采用农杆菌介导的遗传转化方法,将来自枯草杆菌的果聚糖蔗糖转移酶基因(SacB)导入美丽胡枝子,以提高胡枝子抵御干旱胁迫和盐胁迫的能力。以美丽胡枝子子叶节为外植体,通过与含有植物双元表达载体pKP的农杆菌LBA4404共培养,将SacB基因导入美丽胡枝子基因组。经卡那霉素筛选后,共获得62株卡那霉素抗性植株。经PCR特异性扩增和PCR-Southern杂交,证明有5株再生植株基因组DNA中整合了SacB基因。通过RT-PCR分析,结果表明SacB基因均获得表达。经过200mmol/LNaCl和5%PEG模拟胁迫,发现转基因植株美丽胡枝子中,可溶性糖含量在任何时候均高于未转化植株,并比对照拥有更高的抗干旱胁迫和盐胁迫能力。     

番茄转录调控因子基因Pti5植物表达载体的构建及其转化烟草的研究 Construction of a Plant Expression Vector with Tomato Transcription Factor Gene Pti5 and Studies on Transgenic Tobaccos

本实验构建了含有CaMV35S启动子控制下的Pti5-VP16基因的植物双元表达载体pBI121UCH1。通过根癌农杆菌叶盘转化法,将Pti5-VP16基因导入烟草SRI中,经卡那霉素筛选,获得了抗性植株。经PCR和Southern印迹分析,表明抗性植株中整合了Pti5-VP16基因,经抗病性鉴定转基因烟草植株的抗病性明显提高。 Abstract:The plasmid pBI121UCH1 carrying Pti5-VP16 gene under the control of the cauliflower mosaic virus 35s promoter was constructed.Leaf segments of tobacco SRI were infected by Agrobacterium tumefaciens EHA105 with plasmid pBI121UCH1,from which kanamycin resistant plants were obtained.PCR and Southern analysis proved that the Pti5-VP16 gene was integrated into the genomes of the tobacco plants.The disease resistance assay showed that the disease resistance was enhanced in the transgenic tobacco plants.     

提高大岩桐试管苗移栽成活率的实验

取芽或茎尖经HgCl消毒后获得无菌外植体，将其接种于不同组合的培养基上并获得小植株，将小植株进行继代、生根培养，经炼苗后移栽，成活率很高。     

防烟叶霉变菌株对烟叶霉变的影响

为有效防止烟叶霉变,采用平板对峙培养的方法,就3个防烟叶霉变菌株对霉变菌的抑制作用进行了研究．结果表明,JMBl42、B112、B329对供试霉变菌皆表现出一定的抑制作用,对不同霉变菌的平均抑制率分别为51．6％、52．9％、53．7％．3个防烟叶霉变菌株分别以每mL 10^7、10^8、10^9 cfu(colony当当forming unit)13个浓度单独处理和每mL10。cfu浓度混合处理烟叶,结果表明,JMBl42菌株每mL10,cfu处理浓度效果最好,与对照差异极显著,其次为B112菌株每mL10^8cfu处理浓度,但它与对照差异不显著,B329菌株处理效果最差,混合施用结果与对照差异不显著．由此确定JMBl42菌株每mL10^9cfu处理浓度为仓储烟叶防霉变最佳处理参数、