含双病原物诱导启动子无选择标记转基因烟草的获得

Transgenic plants are controversial for their edible and environmental security. Marker-free transgenic plants can be produced by the construction and transformation of plant expression vectors carrying twin T-DNAs. The construction of plant expression vectors harboring twin T-DNAs and two pathogen-inducible promoters was previously reported. These vector plasmids were introduced into tobacco plants and the transgenic tobacco plants were obtained. In this paper we report that T₁ transgenic tobacco seedlings were produced through classical genetics approach. Analysis of the seedlings demonstrated that some of them were marker-free lines. The segregation of exogenous genes in T₁ transgenic tobacco seedlings was tested by using two methods. Firstly, the ability of resistance to kanamycin was analyzed in T₁ transgenic tobacco seedlings from 14 transgenic plant lines. It was found that the segregation ratio of NPTII genes met well with Mendel’s law in 13 transgenic tobacco lines. So it was deduced that NPTII gene was as a single copy integrated into one of the homologous chromosomes. Then the NPTII genes and uidA genes of 130 T₁ transgenic seedlings were detected from the above 13 tobacco lines. The results showed that uidA genes were only detected in 20.77% of the seedlings, NPTII genes were solely detected in 22.31% of the seedlings, but both exogenous genes were in 53.85% of the seedlings. The segregation ratio of the two genes was consistent with the law of independent assortment (9∶3∶3∶1). These results suggested that the selective marker gene had no linkage with the reporter gene and they were segregated independently in the T₁ transgenic tobacco plants. This method, as compared with traditional backcross, is confirmed a more easy and rapid way to eliminate the antibiotic resistance gene used as a selective marker in transgenic plants.     

Elicitin诱导烟草微敏反应和防卫基因的表达

 研究了一种α型elicitin-parasiticein对烟草微敏反应(microscopic hypersensitive response, micro-HR)和防卫反应分子表征基因表达的诱导作用。用150 nmol/L的parasiticein喷洒烟草叶片12 h后, 经曲利本蓝(trypan blue)染色, 光镜下可观察到有细胞坏死, 说明parasiticein引起了micro-HR, 而水和低浓度的parasiticein不能引起micro-HR。用parasiticein注射叶片可引起肉眼可见的HR, 注射用parasiticein浓度远比喷洒引起micro-HR的浓度低。Parasiticein还可诱导对TMV的抗性, 浓度在30 nmol/L时比150 nmol/L的效果好。用parasiticein注射烟草30 min, HR表征基因hsr203J和hin1开始表达, 在9 h内逐渐降低, 12~16 h肉眼可观察到HR。Parasiticein喷洒烟草后, 病程相关蛋白(pathogenesis-related, PR)基因PR-1b在诱导的第1 d到第5 d都有一定量的表达。可见, parasiticein能同步诱导过敏反 应和抗病 性及其分 子表征基 因的表达 。     

烟草赤星病菌糖蛋白激发子诱导烟草抗病防卫反应

用来自赤星病菌弱毒株的糖蛋白激发子处理烟草，能明显抑制烟煤草体内过氧化氢酶（CAT）的活性，增加组织中H2O2的含量，诱导后1dCAT活性就已开始下降，至第6天下降到最低，以后又逐渐恢复，而H2O2含量的变化趋势与CAT活性的变化相反，随CAT活性的降低，组织中H2O2含量逐渐增加，在诱导后第10天达到最大值，激发子处理烟草还可以诱导碱性PR蛋白基因PR1b的表达；同时，烟草对赤星病强毒株的抗性逐渐增强，以诱导后第10天接种抗性最强，且这种抗性是系统性的，水杨酸（SA）处理后CAT及H2O2的变化与上述趋势类似。     

烟草抗赤星病诱导剂SRS2的田间应用

作者在温室、田间上区和大田进行应用示范试验,明确了诱导剂ＳＢＳ２对烟草赤星病的控制作用。室内测定结果表明,ＳＲＳ２与菌核净防治效果相当,均接近１００％。１９９４年在山东沂南作了小区试验,１９９４、１９９５年在山东和安徽８县（市）进行了应用示范试验,结果表明,诱导剂ＳＲＳ２能够较好地控制烟草赤星病,最高可达８５％,平均防治效果为６２．７％,与菌核净相比,ＳＲＳ２的防治效果平均高出２．７％。     

AT毒素胁迫的再生烟草及后代对赤星病的抗性

从4个烤烟品种幼苗心叶叶碟产生的愈伤组织(CA)在含AT毒素的培养基上经两轮筛选,获得的抗毒素CA用于产生了抗病的一次再生植株。用来自抗病性提高50％以上的一次再生株的二代CA产生了抗病的二次再生植株,品种NC89的二次再生抗病植株被称为NC89—TT。获得种子后,NC89—TT二代苗显示出抗毒素损伤、抗病菌侵入、抗病斑扩展的能力,与NC89比,毒素引起的根和叶部病变至少晚8小时、轻64％,病菌孢子侵染率低60％以上,病斑面积减小92％。     

利用GICA-PCR快速检测瓜类果斑病菌

瓜类果斑病菌(Acidovorax avenae subsp.citrulli,Aac)是瓜类作物上重要的病原细菌,为我国进境植物检疫性有害生物。胶体金免疫层析试纸条方便快捷,应用广泛。该方法使用不当会出现假阳性问题,仅适用于病原菌的初筛检测。本研究将Aac胶体金免疫层析方法(GICA)与PCR技术相结合,建立了GICA-PCR检测方法。检测结果表明,该方法在蛋白与核酸2个层面上从发病西瓜叶片上检测到瓜类果斑病菌,有效解决了试纸条检测的假阳性问题,提高了瓜类果斑病菌检测的准确性,值得推广应用。     

小麦纹枯病抗性研究进展

本文主要综述了我国小麦纹枯病病原菌、病症、发病特点、抗性鉴定方法、小麦品种（品系）对纹枯病抗性等方面研究的进展。并从小麦遗传育种角度就今后研究提出了看法和建议。     

生姜茎基腐病病原拮抗细菌的筛选与鉴定

从生姜根际土壤分离得到62个细菌分离物。以姜茎基腐病病原菌(Pythium myriotylum)为指示菌,共选出7株对姜茎基腐病菌有良好拮抗效果的细菌菌株。经过形态学观察、生理生化测定,16S rDNA序列及系统发育分析,结合《常见细菌系统鉴定手册》等,X-11、JK-14、SF-19、SF-23等4株初步鉴定为芽孢杆菌属(Bacillus spp.),JK-19、JK-28、JK-25 3株鉴定为类芽孢杆菌属(Peanibacillus spp.)。SF-19、SF-23、JK-25、JK-19、JK-28、X-11和JK-14的16S rDNA序列的GenBank登录号分别为JQ283970、JQ283971、JQ283972、JQ283973、JQ283974、JQ283975、JQ283976和JQ283977。进一步分析表明,7株拮抗菌对Rhizoctonia solani、Fusarium graminearum、Verticillium dahliae等多种类型的病原真菌有较好的抑制作用;显微观察结果表明,拮抗菌可导致病菌菌丝细胞顶端膨大、菌丝体畸形、分隔增多等变化。     

我国部分地区玉米丝核菌组成及其致病类型分析

IA为主要融合群;双核丝核菌为AG-A融合群;单核丝核菌种类尚不确定.对各融合群的致病类型进行初步比较发现,属于AG1-IA融合群的菌株,可在玉米叶鞘形成典型的云纹状病斑,其它菌株虽可引起玉米发病,但与AG1-IA的症状存在明显差异.     

核黄素诱导烟草悬浮细胞酚类物质和木质素的积累

 【目的】阐明核黄素对烟草悬浮细胞苯丙氨酸解氨酶(phenylalanine ammonia-lyase,PAL)、过氧化物酶(peroxidase,POD)活性以及抗病相关的酚类物质、木质素等次生代谢产物含量的影响。【方法】1 mmol?L-1的核黄素处理烟草悬浮细胞,用生物化学、细胞化学和高效液相色谱(high performance liquid chromatography,HPLC)等方法测定苯丙烷代谢有关的生理和生化指标。【结果】核黄素处理后,PAL和POD活性明显升高,处理12 h 和24 h后酶活性分别达到峰值;荧光观察细胞壁积累了更多的酚类物质,胞内和胞外scopoletin(7-羟基-6-甲氧基香豆素)含量明显升高,在处理12 h后都达到峰值,分别是对照的3.6和3.9倍;细胞染色和含量测定表明,核黄素处理后烟草悬浮细胞沉积了更多的木质素,处理24 h和48 h后,木质素含量分别为对照的1.5和1.8倍。【结论】核黄素处理后提高了烟草悬浮细胞PAL、POD活性,积累了更多与抗病有关的次生代谢产物。