离体筛选抗枯萎病辣椒新种质

以2个辣椒(Capsicum annuum L.)自交系子叶诱导产生的愈伤组织为材料,辣椒枯萎菌粗毒素为选择剂,筛选抗枯萎病辣椒新种质.结果表明,枯萎菌粗毒素对辣椒子叶愈伤组织诱导、生长及不定芽分化具有明显的抑制作用,且随着粗毒素浓度的增加,抑制作用增强;在枯萎菌粗毒素质量浓度为0.60 g·L-1条件下,筛选、鉴定并获得了抗枯萎病辣椒体细胞变异无性系,且成功再生抗性植株.从而证明,离体筛选抗枯萎病辣椒新种质的方法是可行的.     

禾谷镰刀菌粗毒素对不同抗性小麦品种成熟胚脱分化与再分化的影响

以5个赤霉病抗性水平不同的小麦品种为材料,研究了禾谷镰刀菌粗毒素对小麦成熟胚脱分化与再分化的影响.结果表明,小麦成熟胚愈伤组织的诱导率和分化率在品种间、毒素浓度间、毒素浓度×品种间差异极显著.低浓度（5 g/L）禾谷镰刀菌粗毒素能够促进抗性较强品种的出愈速度,高浓度禾谷镰刀菌粗毒素（10 g/L和15 g/L）对成熟胚愈伤组织的出愈速度起到抑制作用,随着浓度提高,抑制作用增强.成熟胚愈伤组织的诱导率和分化率随着禾谷镰刀菌粗毒素浓度升高而下降,品种的抗性越差下降幅度越大.10 g/L毒素浓度,5个品种成熟胚愈伤组织的SOD活性均比对照（0 g/L）增加;20 g/L毒素浓度,5个品种成熟胚愈伤组织的SOD活性均有所下降,抗性品种苏麦3号和郑9023的SOD活性仍高于对照,其它3个感性品种低于对照,SOD活性与小麦品种的赤霉病抗性水平存在一定关系.研究结果为小麦赤霉病抗性育种提供了一定的技术支撑.     

硝酸银对马铃薯叶片组织培养中芽再生的促进作用

硝酸银在马铃薯品种Ｋ４和Ｄｅｓｉｒｅｅ叶片组织培养的愈伤诱导阶段和芽再生阶段中能显著地提高芽再生频率。尤其在愈伤诱导阶段中它可刺激芽的直接形成而不经过愈伤中愈伤诱导和芽再生作用没有硝酸银显著,亦不能促进愈伤诱导阶段的芽直接形成。２,４－Ｄ对愈伤组织形成无明显效果,但可促进愈伤的生长;另外,２,４－Ｄ对芽分化联合会的芽再生具有显著抑制作用,这种抑制作用可被硝酸银所抵消。结果显示在马铃薯叶片组织培养过     

硝酸银对马铃薯叶片组织培养中芽再生的促进作用 (英)

硝酸银在马铃薯品种K4和Desiree叶片组织培养的愈伤诱导阶段和芽再生阶段中能显著地提高芽再生频率。尤其在愈伤诱导阶段中它可刺激芽的直接形成而不经过愈伤形成过程。硫代硫酸银对叶片组织培养中愈伤诱导和芽再生作用没有硝酸银显著,亦不能促进愈伤诱导阶段的芽直接形成。2,4－D对愈伤组织形成无明显效果,但可促进愈伤的生长;另外,2,4－D对芽分化阶段的芽再生具有显著抑制作用,这种抑制作用可被硝酸银所抵消。结果显示在马铃薯叶片组织培养过程中具有乙烯的生物合成,通过调节乙烯的合成或作用过程可以改良马铃薯叶片的组织培养效果。     

马铃薯无菌苗叶肉原生质体再生植株

商用马铃薯叶肉原生质体,在不同的液体培养基中浅层培养,耳生细胞分裂,并获得愈伤组织。将愈伤组织转到固体培养基上诱导分化,已从克新四号和68—62两个马铃薯品种的原生质体得到再生的完整植株。再生细胞的分裂频率受原生质体培养密度的影响。细胞团的生氏对液体培养基中的蔗糖浓度敏感。不同的原生质体培养基对愈伤组织的分化频率的影响非常显著。在分比培养基中加入3％的甘露醇,能提高愈伤组织的分化频率,并缩短分化期。     

马铃薯晚疫病抗病基因研究进展

马铃薯晚疫病是马铃薯和番茄等茄科植物中最主要的病害之一,每年都引起巨大的经济损失。基因工程技术的发展为马铃薯晚疫病的防治工作提出了新的契机,从晚疫病抗性品种中筛选出具有高持久抗性的抗病基因,并将其转化到栽培品种中去,无疑是我们开发持久性晚疫病抗性的最快捷的手段。到目前为至,已经有十几个晚疫病抗性基因从S．demissum,S. bulbocastanum,S．berthauhii,S．mochiquense和S．pinnatisectum等抗性马铃薯品种中鉴定出来并已定位在马铃薯染色体基因组上,并有4个被克隆出来（R1,R3a,Rpi—blb1／RB和Rpi—blb2）。主要概述了马铃薯晚疫病抗病基因的研究现状和发展前景。     

利用水稻花药培养筛选稻瘟病抗性突变体

近年来,利用植物组织培养的方法并结合使用抗源物粗毒素来筛选抗病突变系,已在不同物种中获得了成功。例如,Ingram和Robertson利用组织培养研究了马铃薯晚疫病(Phytophthora intesans)的抗性基因表达;Behnke将抗晚疫病毒素的培养细胞进行筛选,成功地获得抗性突变体,     

马铃薯晚疫病抗性基因分子标记检测及抗性评价

马铃薯是重庆优势特色作物之一,但其安全生产受到晚疫病的严重威胁。马铃薯生产中种植抗病品种是防控马铃薯晚疫病最为经济有效和环境友好的途径。为了了解来自国内外不同种质的晚疫病抗性基因组成以及确定在重庆市具有晚疫病抗性的马铃薯品种(系),本研究以218份来自国内外的品种(系)为材料,进行了6个晚疫病抗性(R)基因分子标记检测,同时进行了田间晚疫病抗性评价及室内接种鉴定和筛选。研究结果显示,6个R基因的分子标记在供试材料中均有分布,但分子标记的组成不尽相同,主要分为4大类。第Ⅰ类含有具有广谱抗性基因的RB,晚疫病抗性评价表现为中抗以上;第Ⅲ类缺失R2 family基因标记,绝大部分表现为晚疫病敏感型;第Ⅱ类和第Ⅳ类中分别主要含有3个R基因(R2 Family+R3a+R3b)标记类型和4个R基因(R1+R2 Family+R3a+R3b)标记类型,这两类材料中表现出一定比例的晚疫病抗性水平,但第Ⅳ类中出现抗性表现的比例高于第Ⅱ类。结果说明含有RB基因标记贡献了较高的晚疫病抗性,缺失R2 famlily基因标记的材料可能不利于晚疫病抗性,利用这些基因标记辅助筛选有助于提高重庆地区晚疫病抗性育种效率。本研究评价了218份马铃薯材料6个重要R基因组成,并筛选出重庆地区表现抗性的多个材料,为新品种(系)的推广应用以及抗病育种选育提供了科学依据,同时为发掘新的抗病基因提供了遗传资源。     

马铃薯种质资源的晚疫病抗性鉴定

采用离体叶片接种法,对43份马铃薯种质资源进行晚疫病抗性鉴定、比较和评价。以接种5 d后的感病品种‘Désirée’和抗病品种‘加湘1号’叶片症状为对照,鉴定出5份表现为高抗的种质资源,其中包含3个Solanum phurejia和2个S. tuberosum ssp. andigena材料。另外,还鉴定出14份中抗材料(No. 7～20)。结果表明,野生种和安第斯山栽培亚种马铃薯资源的抗性材料较为丰富,可作为晚疫病抗病育种的亲本。     

利用抑制差减杂交技术分离马铃薯晚疫病抗性相关基因

以晚疫病病原菌混合小种接种处理48h的马铃薯水平抗性材料(R-gene-free)叶片为目的材料,以未处理材料作为对照,用抑制差减杂交技术构建了一个富集晚疫病抗性相关基因的差减文库。应用反向Northern技术对840个克隆进行斑点杂交筛选,筛选出150个病原诱导后信号明显增强的克隆。26个片段测序结果表明：部分片段基因功能与抗病性明显相关。7个差异表达片段与GenBank EST数据库中已有晚疫病原诱导马铃薯叶片得到的EST有很高同源性(达95％～100％);部分片段核苷酸或氨基酸序列分别与番茄、烟草、拟南芥等的EST序列或氨基酸序列有较高同源性;另有4个基因片段在GenBank EST数据库中未找到明显的同源序列,可能为新发现的基因片段。     

Purification of an isoform of patatin with antimicrobial activity against Phytophthora infestans.

Phytophthora infestans (Mont.) de Bary is infamous as the causal agent of the late blight epidemic contributing to the Irish potato famine of the mid 19th century and remains agriculture's most destructive disease as new mutations and migrations confound control measures. In efforts to develop resistant varieties, a somatic hybrid (the Wisconsin J series) between potato (Solanum tuberosum) and a wild relative (Solanum bulbocastanum) has been found to convey durable resistance against the pathogen. We screened the total protein (100 microg ml(-1)) of somatic hybrid varieties J138, J138A12, J101K12, J103K12, and J101K9 for in vitro spore germination inhibition of P. infestans. Since J138 exhibited maximum inhibition at 150 microg ml(-1) in comparison to other varieties, we purified a 40 kD protein from J138 tubers by assaying its ability to inhibit spore germination in P. infestans spores. The highly purified protein was able to inhibit P. infestans spore germination by 70% at the 2.5 microg ml(-1) concentration. The N-terminal sequence of this protein was found to have exact amino acid homology to patatin, the major storage protein of potato tubers. The inhibitory protein has the same molecular weight as patatin and cross-reacts with patatin antibodies. The infection of J138 plants with spores of P. infestans under greenhouse conditions showed that patatin is expressed in stem tissue 72 h after the plant is inoculated with field isolates of P. infestans (US8). In this communication, we report the purification, characterization and antifungal activity against spores of P. infestans of patatin-J from potato tubers.     

Resistance to Phytophthora infestans in somatic hybrids of Solanum nigrum L. and diploid potato

In breeding for resistance to late blight, ( Phytophthora infestans Mont. de Bary), an economically important disease affecting potatoes, the search for new sources of durable resistance includes the non-host wild Solanum species. The aim of this work was to evaluate the resistance to P. infestans in the somatic hybrids between S. nigrum L. and diploid potato clone ZEL-1136. Sixteen somatic hybrids, their fusion parents, and three standard potato cultivars were screened for resistance to P. infestans in two types of tests-on whole plants and detached leaves-with two highly aggressive and virulent isolates of P. infestans, US8 and MP322. In the whole plant assay, the foliage of the somatic hybrids showed no symptoms of infection, while the foliage of the potato fusion parent and the standard cultivars was infected with P. infestans. In the detached leaflet assay, the breaking-down of resistance of the S. nigrum L. parent and the variable response of individual hybrid clones were noted. Nine S. nigrum L. (+) ZEL-1136 hybrids showed a resistance that was significantly higher than that of S. nigrum, while six clones expressed a resistance to P. infestans similar to that of S. nigrum. The results confirm the effective transfer of late blight resistance of S. nigrum into its somatic hybrids with potato.     

马铃薯非共生血红蛋白基因StHb1的克隆及表达

采用RT-PCR技术成功分离了马铃薯StHb1基因序列.经半定量RT-PCR分析表明,StHb1基因的表达在抗性品种(陇薯三号)和感性品种(荷兰十五)块茎中均受致病疫霉的侵染所抑制;StHb1基因在正常生长的马铃薯块茎组织中表达量最高;外源NO和H2O2的作用可明显地抑制StHb1基因的表达,但在抗性品种中该基因受抑制的程度低于感性品种.上述试验结果暗示了StHb1基因与马铃薯对致病疫霉侵染的抗性应答具有一定的相关性.     

Comparative analysis of Phytophthora infestans induced gene expression in potato cultivars with different levels of resistance

Differential gene expression was analyzed after infection with Phytophthora infestans in six potato cultivars with different levels of resistance to late blight. To verify the infection of the potato leaflets, the amount of phytopathogen mRNA within the plant material was quantified by real-time quantitative PCR. The expression of 182 genes selected from two subtracted cDNA libraries was studied with cDNA array hybridization using RNA from non-infected and infected potato leaflets. Gene up- and down-regulation were clearly detectable in all cultivars 72 h post inoculation. Gene expression patterns in susceptible cultivars differed from those in potato varieties with a higher level of resistance. In general, a stronger gene induction was observed in the susceptible cultivars compared to the moderately to highly resistant potato varieties. Five genes with the highest homology to stress and/or defence-related genes were induced specifically in the susceptible cultivars. Four genes responded to pathogen attack independently of the level of resistance of the cultivar used, and three genes were repressed in infected tissue of most cultivars. Even in the absence of P. infestans infection, six genes showed higher expression levels in the somewhat resistant cultivars Bettina and Matilda. Possible reasons for the different levels of gene expression are discussed.     

THE REACTION OF VIRUS-INFECTED POTATO PLANTS TO PHYTOPHTHORA INFESTANS

The growth of Phytophthora infestans was retarded on leaves of potato plants that had been artificially inoculated with virus X or with virus Y.
Using different virus strains and potato varieties, the effect of virus infection on blight development was found to be greater, the more severe the systemic virus symptoms exhibited on the infected leaves before P. infestans inoculation.
The development of the fungus was never increased by virus infection.
The reduced blight development on virus-infected leaves is partially caused by an increase of resistance to infection. It is also suggested that virus infection alters the nutritional status of leaves to one less favourable for the development of P. infestans.     

马铃薯晚疫病生防木霉菌的筛选及鉴定

采用马铃薯活体筛选法从268株木霉菌中筛选获得两株对致病疫霉有较强抑菌活性的木霉菌株R-5和T-15。这两株木霉菌代谢液可抑制病原菌生长及孢子囊萌发。温室防病试验发现,接种两株木霉菌可以减轻晚疫病的发生。田间试验进一步证明,两株木霉菌对晚疫病具有良好的田间防治效果,防效分别达到了72.4%和70.0%。经分子生物学方法鉴定,两株木霉菌分别为拟康氏木霉和棘孢木霉。实验构建的以活体筛选为基础的生防木霉菌筛选方法是一种可行高效的生防木霉菌筛选方式。