Genetic diversity of sulfonylurea-resistant and -susceptible Monochoria vaginalis populations in Japan

Monochoria vaginalis is one of the most serious weeds of rice fields in Asia. The species is predominantly selfing. To reveal the potential for multiple mutational events, outcrossing and gene flow in the sulfonylurea‐resistant (SU‐R) M. vaginalis populations, we investigated (i) if each SU‐R population was a single SU‐R biotype or a mixture of several SU‐R biotypes using restriction analysis or direct sequencing of acetolacatate synthase (ALS) genes and (ii) genetic diversity of SU‐R and ‐susceptible (S) populations using amplified fragment length polymorphism (AFLP) analysis. Nineteen or 20 individuals were sampled from four SU‐R and five SU‐S populations respectively. Amino acid substitutions conferring resistance in the SU‐R populations were Pro₁₉₇Ser in the ALS1 or ALS3, or Asp₃₇₆Glu in the ALS1 and each SU‐R population was composed of a single SU‐R biotype. In cluster analysis each SU‐R individual formed a cluster, whereas the individuals from a SU‐S population belonged to different clusters. Some SU‐R populations showed polymorphic AFLP loci. The results indicated that these SU‐R biotypes emerged from a single mutational event and any gene flow of SU‐R genes from adjacent populations did not occur. A low level of outcrossing and recombinations of SU‐R genes occurred within some SU‐R populations of M. vaginalis.     

Relation between pathogenicity and genetic variation within <Emphasis Type="Italic">Plasmodiophora brassicae</Emphasis>

The relation between diversity of pathogenicity on clubroot-resistant (CR) cultivars of Chinese cabbage (Brassica rapa subsp. pekinensis) bred in Japan and DNA polymorphisms in 17 populations of Plasmodiophora brassicae from cruciferous plants was examined by inoculation tests and random amplified polymorphic DNA (RAPD) analysis using 18 arbitrary primers. Four pathotypes (A–D) were identified after inoculation of six CR cultivars of Chinese cabbage in the 17 populations from cruciferous crops. A relatively high level of genetic diversity was also detected among these populations in the RAPD analysis. Although the four pathotypes could not be clearly differentiated using the RAPD data, most populations of three pathotypes had a consistent location on the dendrogram. All pathotype B (virulent on five cultivars except Utage 70) and D (avirulent on all cultivars) populations, which were common in incompatible interactions with cv. Utage 70, were located in a single subcluster. All five pathotype C populations (virulent only on cv. Utage 70) except for one population grouped in another single subcluster. Because four pathotype A populations (virulent on all six cultivars, races 4 and 9) fell in different subclusters, the populations may be genetically polyphyletic. Populations from cruciferous weed Cardamine flexuosa differed remarkably from those from cruciferous crops in pathogenicity on common cultivars of Chinese cabbage and turnip and C. flexuosa, but they grouped in a single cluster with all race 9 populations from crops. Race 9 populations from crops may thus be closely related to populations from the weed rather than to races 1 and 4 from crops.     

Genetic structuring and diversity patterns along rivers – local invasion history of Ambrosia artemisiifolia (Asteraceae) along the Danube River in Vienna (Austria) shows non‐linear pattern

Ambrosia artemisiifolia is an annual weed from North America that nowadays is invasive in many countries worldwide. In Austria, numerous populations of A. artemisiifolia are located along the Danube River, especially along the ‘New Danube’ (Vienna). This area is characterised by ruderal and riparian sites, which are regularly flooded. To better understand the spread of A. artemisiifolia and its colonising behaviour along the Danube River, we analysed genetic structure and diversity based on 23 populations linearly arranged along the Viennese Danube riverbed and upstream, utilising the Amplified Fragment Length Polymorphism (AFLP) fingerprint method. We generated 284 polymorphic AFLP markers across 446 A. artemisiifolia plants. The genetic diversity within populations was higher (HW = 0.091) than among populations (HB = 0.007). This result indicates A. artemisiifolia introductions from similar mixtures of sources or spread from a single already mixed introduction. Within our local setting, we were unable to identify neither source or sink populations nor an obvious linear genetic structuring. Genetic among‐population differentiation was low to moderate (amova ‐derived F_ST = 0.124). Lack of geographical structuring is indicative of highly dynamic gene flow, which is further supported by the absence of an isolation‐by‐distance pattern. Multiple introductions and non‐directional gene flow are most likely promoted by anthropogenic disturbance and human‐mediated dispersal. Our results demonstrate the ability and speed of A. artemisiifolia to settle in newly disturbed areas and the difficulties to predict invasion directions, as downstream river dispersal was negligible.     

小菜蛾对溴氢菊酯抗药性的随机扩增多态性DNA分析

利用室内选育119代的抗溴氰菊酯小菜蛾Plutella xylostella品系和敏感品系，通过反复回交和自交建立近等基因系。用Operon公司合成的20个引物对近等基因系基因组DNA进行PCR扩增。3个引物在抗性品系中分别产生1条特异扩增带，4个引物在敏感品系中分别产生了1～5条特异扩增带。由于在近等基因系中，除了与抗性相关的基因区外，其它的遗传背景是相同的，因此可以认为这些特异带与小菜蛾对溴氰菊酯的抗性有关。     

Genetic diversity of the root‐knot nematode Meloidogyne ethiopica and development of a species‐specific SCAR marker for its diagnosis

Meloidogyne ethiopica is an important nematode pathogen causing serious economic damage to grapevine in Chile. In Brazil, M. ethiopica has been detected with low frequency in kiwifruit and other crops. The objectives of this study were to evaluate the intraspecific genetic variability of M. ethiopica isolates from Brazil and Chile using AFLP and RAPD markers and to develop a species‐specific SCAR‐PCR assay for its diagnosis. Fourteen isolates were obtained from different geographic regions or host plants. Three isolates of an undescribed Meloidogyne species and one isolate of M. ethiopica from Kenya were included in the analysis. The results showed a low level of diversity among the M. ethiopica isolates, regardless of their geographical distribution or host plant origin. The three isolates of Meloidogyne sp. showed a high homogeneity and clustered separately from M. ethiopica (100% bootstrap). RAPD screenings of M. ethiopica allowed the identification of a differential DNA fragment that was converted into a SCAR marker. Using genomic DNA from pooled nematodes as a template, PCR amplification with primers designed from this species‐specific SCAR produced a fragment of 350 bp in all 14 isolates of M. ethiopica tested, in contrast with other species tested. This primer pair also allowed successful amplification of DNA from single nematodes, either juveniles or females and when used in multiplex PCR reactions containing mixtures of other root‐knot nematode species, thus showing the sensitivity of the assay. Therefore, the method developed here has potential for application in routine diagnostic procedures.     

Development of sequence tagged site markers to identify races of Fusarium oxysporum f. sp. lactucae

By random amplified polymorphic DNA (RAPD) analysis of the representative isolates of each race of Fusarium oxysporum f. sp. lactucae, RAPD fragments of 0.6, 1.6, and 2.9kb were obtained. The 0.6-kb RAPD fragment was common to the representative isolates of all three races. Amplification of the 1.6- and 2.9-kb fragments were unique to the isolates of races 1 and 2, respectively. Sequence tagged site (STS) marker FLA0001, FLA0101, and FLA0201 were generated from the 0.6-, 1.6-, and 2.9-kb RAPD fragments, respectively. Polymerase chain reaction (PCR) analysis showed that FLA0001 was common to all 49 isolates of F. oxysporum f. sp. lactucae. FLA0101 was specifically generated from all 23 isolates of race 1 but not from races 2 or 3. FLA0201 was specifically amplified from all 12 isolates of race 2 but not from races 1 or 3. In two isolates of F. oxysporum f. sp. lactucum, PCR amplified FLA0001 and FLA0101 but not FLA0201. On the other hand, these STS markers were not detected from isolates of five other formae speciales. Because these STS markers were not generated from isolates of other plant pathogenic fungi, bacteria, or plant materials examined in this study, PCR analysis combined with the three STS markers should be a useful means for rapid identification of races of F. oxysporum f. sp. lactucae.     

Random amplified polymorphic DNA markers reveal low genetic variation and a single dominant genotype in Eichhornia crassipes populations throughout China

Genetic variation within and between 34 populations of Eichhornia crassipes (water hyacinth) in China was surveyed using random amplified polymorphic DNA (RAPD) markers. A total of 1009 individuals were analysed, for which 12 RAPD primers amplified 69 reproducible bands, with 22 (32%) being polymorphic. The percentage of polymorphic loci (p) within a population ranged from 4.4% to 17.4%, and the mean Nei's gene diversity (H_e) was 0.046 ± 0.0145, indicating a low genetic diversity of E. crassipes in China. Each population contained at least four RAPD phenotypes (genotypes), and the same particular genotype was invariably dominant in all the populations sampled. The mean proportion of distinguishable genotypes was 0.29. Analysis of molecular variance revealed a large proportion of genetic variation (83.9%) residing within populations and a slightly larger proportion (88.1%) within localities, indicating a low genetic differentiation of E. crassipes populations, both locally and regionally. Human-mediated dispersal, vigorous clonal growth, and a generally low level of sexual reproduction were thought to be responsible for such a pattern of genetic structure.     

福建若干荔枝古树资源的RAPD分析

利用随机扩增多态性DNA(RAPD)技术,从104个十聚体随机引物筛选出13个引物对福建荔枝著名的古树以及相关品种等12份材料的基因组DNA进行扩增,共得到78条扩增谱带,其中2条为共同带,多态性程度达97.44%,平均每条引物扩增的谱带数目为6条.采用遗传标记计算遗传资源相似性系数,进行聚类分析,并构建树状分析图.结果表明:现在栽培的陈紫品种与宋家香的亲缘很近,可能来源于宋家香;而元红与西禅寺“宋荔”亲缘关系较远.应用RAPD技术为荔枝古树遗传资源的鉴定和分类提供了新的途径.     

红富士苹果芽变系DNA甲基化研究

以35份富士苹果(Malus×domestica Borkh.‘Fuji’)芽变材料为试材,利用甲基化敏感扩增多态性(Methylation Sensitive Amplified Polymorphism,MSAP)分析和UPGMA聚类方法,对其基因组甲基化修饰水平、变异模式以及表观遗传变异关系进行研究。结果表明:(1)不同富士系得到不同的MSAP扩增,总DNA甲基化水平27.90%～36.16%,平均32.87%,双链全甲基化为主要甲基化方式;(2)富士芽变材料绝大多数位点保持了原有甲基化模式;(3)绝大多数芽变(68.57%)检测到全部的甲基化变异模式(12种),去甲基化频率极显著高于甲基化频率(P <0.01),且CG去甲基化极显著高于CHG;(4)36份种质遗传相似系数平均值0.89(0.79～0.92),在聚类图上,富士原种分布在芽变系集中区外,新近发生的芽变系更倾向于聚在一起,着色系片红型和条红型芽变呈分散排布状态。总的来看,富士芽变的甲基化变异模式丰富,超甲基化和去甲基化相伴发生,但以去甲基化为主;‘富士’着色芽变与其最原始品种富士,以及芽变之间发生了较大表观遗传变异;片红和条红型芽变聚类未表现明显偏好性。本研究将为进一步开展富士着色系芽变机理研究提供指导,可以CG去甲基化为切入点展开深入研究。     

利用SRAP分子标记技术鉴定大白菜种子纯度

以大白菜品种多抗3及其父母本为研究材料,利用SRAP（相关序列扩增多态性）分子标记方法,通过对54对SRAP标准引物的筛选,获得了能区分大白菜多抗3及其亲本的引物Me4F/Em3R。采Me4F/Em3R引物对对5份多抗3系列商品种子纯度进行检验,种子的纯度分别为75.5%、72%、80.5%、68%和73%,与田间种植...