Molecular characterization of chestnut plants selected for putative resistance to Phytophthora cinnamomi using SSR markers

A set of nine short sequence repeat (SSR) loci was used for the molecular characterization of 32 accessions of 15 chestnut trees selected in the field because of their putative resistance to the ink disease caused by Phytophthora spp. The goal of the present study was to determine the genetic identity of those selected European chestnut trees (Castanea sativa) or interspecific hybrids, considering that hybridization programs between European chestnut and Asiatic species (mainly Japanese chestnut, Castanea crenata) have been carried out in Galicia (Spain) since the early 20th century. The results showed that the analyzed SSR loci were useful to discriminate three Asiatic and the European species of Castanea. The joint information provided by a factorial correspondence analysis (FCA) and the presence of privative alleles allowed the putative molecular assignment of the selected plants to a certain identity. Most of them were determined as hybrids between C. crenata and C. sativa. The individuals coded C036 and C048 were assigned, with a high probability, to C. sativa due to their clustering with accessions of this species and because they had a number of privative alleles of this species. Only a few individuals could not be assigned to any particular genotype.     

Genetic diversity, aggressiveness and metalaxyl sensitivity of Pythium aphanidermatum populations infecting cucumber in Oman

Seventy three isolates of Pythium aphanidermatum obtained from cucumber from four different regions of Oman and 16 isolates of muskmelon from the Batinah region in Oman were characterized for aggressiveness, sensitivity to metalaxyl and genetic diversity using AFLP fingerprinting. Twenty isolates of P. aphanidermatum from diverse hosts from different countries were also included in the study. Most isolates from Oman were found to be aggressive on cucumber seedlings and all were highly sensitive to metalaxyl (EC₅₀ < 0·80 µg mL^?1). Isolates from cucumber and muskmelon were as aggressive as each other on both hosts (P > 0·05), which implies a lack of host specialization in P. aphanidermatum on these two hosts in Oman. AFLP analysis of all isolates using four primer–pair combinations resolved 152 bands, of which 61 (~40%) were polymorphic. Isolates of P. aphanidermatum from Oman and other countries exhibited high genetic similarity (mean = 94·1%) and produced 59 different AFLP profiles. Analysis of molecular variance indicated that most AFLP variation among populations of P. aphanidermatum in Oman was associated with geographical regions (F_ST = 0·118; P < 0·0001), not hosts (F_ST = –0·004; P = 0·4323). These data were supported by the high rate of recovery (24%) of identical phenotypes between cucumber and muskmelon fields in the same region as compared to the low recovery (10%) across regions in Oman, which suggests more frequent movement of Pythium inoculum among muskmelon and cucumber fields in the same region compared to movement across geographically separated regions. However, recovering clones among regions and different countries may imply circulation of Pythium inoculum via common sources in Oman and also intercontinental spread of isolates.     

Comparative proteomic analysis of Orobanche and Phelipanche species inferred from seed proteins

Orobanche and Phelipanche species are root parasitic plants some of which infect important crops, causing extensive damage. In an attempt to unravel molecular systematic in these genera using protein markers, we established the protein profile of twelve species by two-dimensional gel electrophoresis (2-DE). Proteins were detected by Sypro Ruby staining and the gel images were digitalised and analysed. The 2-DE maps contained from 30 to 140 protein spots for the analysed species. A virtual master gel was built with spots present in all species and revealing the presence of 260 protein spots. Only one protein was monomorphic, while 143 were unique to a single species. Proteins were scored for the presence or absence of homologous spots to create a binary matrix. Phylogenetic analysis was carried out using maximum parsimony and distance method. The trees obtained were rooted using Cistanche phelypaea as an out-group. Both analyses showed Orobanche as a monophyletic group. AMOVA analysis revealed phenotypic diversity between species within genera, as well as a considerable level of variation between genera. This is a first step to help in the phylogenetic classification of parasitic plants by means of proteomics.     

RAPD and ITS-based variability revealed in Atriplex species introduced to semi-arid zones of Morocco

Variability related to RAPD markers and sequences of nrDNA ITS region has been studied in seven species of the genus Atriplex (A. amnicola, A. canescens, A. halimus from Morocco (MAR) and from USA, A. lentiformis, A. nummularia, A. semibaccata and A. undulate). As a whole, the results show a high variability among the species. The biggest diversity was obtained by RAPD data, followed by ITSs. According to RAPD markers, two major groups can be distinguished, one formed by A. semibaccata and A. undulata, the least similar to the rest of species. A. lentiformis was closer to A. halimus. Phylogenetic analysis confirmed the divergence of A. semibaccata from all species and the closeness of A. lentiformis to A. halimus. The intraspecies variability was also high, as 39 individual RAPD haplotypes were obtained by analyzing 40 plants. Genetic diversity was bigger among species (60.23%) than among individuals (39.77%). The amplification of ITS region leads to three well-defined clades. The heterogeneous and larger clade includes all A. amnicola individuals and some individuals from A. nummularia, A. lentiformis, A. halimus USA, A. halilmus MAR and A. undulata. Both RAPDs and ITS analyses revealed with some exceptions, that all individuals from one species grouped together. RAPDS turn out to be more appropriate than ITS to differentiate Atriplex species. The two markers gave rise to the same species relationships, but to a different structure of the Atriplex genetic diversity. AMOVA analysis estimated interspecies differences for 11.20% of the total ITS variation.     

Analysis of genetic diversity in Turkish sesame (Sesamum indicum L.) populations using RAPD markers⋆

The genetic diversity of 38 cultivated populations of Sesamum indicum L. from four different regions of Turkey was estimated at the DNA level with the random amplified polymorphic DNA (RAPD) technique. Sixty-one bands were obtained for all populations 78% of which were polymorphic. Analysis of molecular variance (AMOVA) was used to investigate the genetic diversity of the populations which yielded highly significant differences among populations within regions (91.9% of the total genetic diversity). According to AMOVA and Shannon's index that were performed separately for each region, the highest value of genetic variation was observed among Northwest region populations (CV = 7.7; H₀ = 0.304) and lowest in the Southeast regions' populations (CV = 2.6; H₀ = 0.068). Nei and Li's similarity index was calculated and phylogenetic tree was established using the neighbor-joining algorithm. This phenetic analysis grouped 35 of 38 accessions in six groups leaving three highly diverse accessions outside. Wagner phylogenetic method was used to assess the phylogenetic relationships among the populations. In the majority-rule consensus tree, only 7 of the 32 forks showed above 60% occurrence. Using Principal Coordinate Analysis (PCO) of the RAPD data set, the groups were clearly separated along the first three axis. These results indicate that RAPD technique is useful for sesame systematics, and should be valuable for the maintenance of germplasm banks and the efficient choice of parents in breeding programs.     

利用RAPD-PCR与ISSR-PCR标记技术分析长江口刀鲚的群体遗传结构

利用RAPD-PCR及ISSR-PCR两种分子标记技术对长江出海口两年3个群体的52条刀鲚（Goiliaectenes）进行群体遗传结构的分析。在3个群体中，利用RAPD-PCR标记技术，15个10bp随机引物共检测到110条带，多态性为0．490～0．657，Shannon多样性指数为18．63～22．38，Nei平均遗传距离为0．1195～0．1454；而利用11个ISSR引物所获得的相应结果分别为67、0．576～0．682、11．56～13．66以及0．117～0．147。相关性分析表明这两种技术所获得的上述数据呈正相关（r=0．95，P〈0．05）。AMOVA结果显示，3个群体按采集时间划分成的两年之间的遗传变异不超过总变异的1．1％，同一年内群体间的遗传变异也分别只占总变异的6．99％（RAPD）和2．75％（ISSR-PCR），群体内各个体之间的遗传变异占总变异的96％（P〈0．0001），说明两年内所采集的3个群体之间基本上没有产生遗传分化。基于样品之间的Nei遗传距离所构建的Neighborjohing聚类图也清楚地显示出没有按采样的时间产生群体的遗传分化。对各样品之间的Nei遗传距离及Neighbor-joining聚类图分别经Mantel检测及支序分析，发现ISSR-PCR技术所获得的结果与RAPD-PCR技术的结果不存在明显的正相关，但ISSR-PCR标记技术具有在待测样品中能检测到高多态性等优点。     

Evidence of strong gene flow among French populations of the carrot cyst nematode Heterodera carotae

The recent ban of the most efficient chemical nematicides has left growers without methods for controlling the carrot cyst nematode Heterodera carotae. This phytoparasitic nematode species has a very narrow host range and causes severe crop losses in the main carrot-growing regions worldwide. The development of alternative means of management of H. carotae is thus essential, and knowledge is required about the adaptive abilities of H. carotae, which mainly depend on gene flow among populations. The goal of this study was to describe the genetic structure of H. carotae populations at the spatial scale of the main infested French carrot-producing region, i.e. Lower Normandy, and to disentangle the causes of the heterozygote deficit in this polyvoltine species. Microsatellite genotyping of populations collected at both the plant and field scales showed that: (i) the heterozygote deficit is mainly due to substructure; and (ii) there is strong gene flow among populations, leading to low F_ST and to no clear genetic structure at the spatial scale explored here. Soil transport through both agricultural machinery and the transport of leek seedlings is probably responsible for the very strong H. carotae migration among fields and production areas. Measures should be considered to limit the passive spread of H. carotae.     

洪泽湖河蚬(Corbicula fluminea) 2种表型群体的遗传变异分析

采用形态学分析(壳长、壳宽和壳高)和分子标记技术(细胞色素氧化酶Ⅰ基因,COⅠ)对洪泽湖河蚬(Corbicula fluminea)黄色和黑色2个群体的形态和遗传多样性特征进行研究.形态参数统计分析结果显示,2个群体的形态特征之间有显著性差异.经PCR扩增和序列测定,获得614 bp COⅠ基因序列,2个群体的COⅠ基因序列的碱基组成高度一致,均表现出A+T的含量(64.8％)明显高于G+C的含量(35.2％).28个黑色个体发现7种单倍型,单倍型多样性和核苷酸多样性分别为0.794和0.04274;30个黄色个体发现5种单倍型,单倍型多样性和核苷酸多样性分别为0.607和0.02825.单倍型之间的遗传距离在0.002-0.091之间,其NJ和MP系统发生树表明,COⅠ基因单倍型聚为2个明显分支.分子方差分析(AMOVA)结果显示,Fst=0.21736 (P＜0.01),21.74％的变异来自群体间,78.26％的变异来自群体内,2个群体之间有显著的遗传分化.研究表明,应将洪泽湖河蚬黑色和黄色群体分别作为独立单元进行管理和保护.     

Genetic diversity in cornsalad (Valerianella locusta) and related species as determined by AFLP markers

Fifteen amplified fragment length polymorphism (AFLP) EcoRI/MseI‐based primer combinations with five selective bases (Eco RI‐ANN, MseI‐CN) were used to estimate genetic diversity among 45 line varieties of cultivated cornsalad and 19 genebank accessions classified into nine different species related to cornsalad. Polymorphic fragments were scored for calculation of Jaccard's coefficient of genetic similarity (GS). The average GS estimate in elite germplasm (GS = 0.90) was substantially higher than in exotic germplasm (GS = 0.47). UPGMA‐cluster analysis revealed genetic relationships among recently bred varieties, old varieties and genebank accessions. Analysis of molecular variance indicated almost threefold variability within sets compared with between sets due to a high level of polymorphism among wild species. Sources for increasing genetic diversity in elite germplasm of cornsalad were suggested and a duplicate among the genebank accessions was detected. AFLPs could be considered a powerful tool for genetic diversity estimation in cornsalad germplasm and are recommended for systematic fingerprinting of remaining cornsalad species.     

中国节节麦基于ISSR标记的遗传多样性分析

为揭示中国节节麦的遗传多样性并发掘可能具有独特变异的节节麦种质资源,采用ISSR标记对75份中国节节麦的遗传多样性进行分析。结果表明,筛选出的9条ISSR引物共检测得到155个位点,多态性位点(138个)百分率为89.31%。Nei’s多样性指数(He)、Shannon’s信息指数(I)、基因分化系数(Gst)和基因流(Nm)分别为0.273 4、0.415 2、0.138 5和3.11,表明中国节节麦有较高的遗传多样性,群体间有中等程度的遗传分化。基于简单相似系数的UPGMA聚类分析表明,除5份河南和陕西节节麦聚类形成独立的分支Group 2(T078、T102和SC1)和Group 3(SX38和T006)外,绝大多数节节麦均聚类形成较大的分支Group 1,且在Group 1中,除4份节节麦(T002、T023、XJ6和XJ49)外,绝大多数节节麦均依据地理分布分别形成了黄河流域节节麦亚组和新疆节节麦亚组,在遗传距离约0.77处,又可依次分为河南、陕西和新疆节节麦小分支。二维主成分分析、群体的遗传一致度和分子变异分析也表明了类似的结果,同属于黄河流域节节麦亚组的河南、陕西节节麦遗传一致度(S=0.971 8)较高,群体内个体差异是中国节节麦变异的主要原因,但两大亚组和三居群的遗传差异分别占总变异的18.57%和10.38%。可见,不同节节麦的地理分布和生境差异,是导致中国节节麦居群遗传分化的主要原因,而聚类分析中单独形成独立分支Group 2和Group 3的5份黄河流域节节麦,可能是具有独特遗传变异的种质资源。