青藏高原青稞淀粉资源的评价

青稞（Hordeum vulgare L.var．nudum Hook．f.），即裸大麦，是兼食用、饲用和酿造于一体的作物，有着重要的利用价值。淀粉是青稞籽粒中含量最多、最重要的碳水化合物，淀粉含量、直支淀粉比将会直接影响淀粉的功能特性，进而影响淀粉的应用领域。我国青藏高原青稞的栽培和食用历史悠久，特色青稞资源极其丰富。目前关于青藏高原青稞淀粉特性的报道还不多见，筛选和培育特色淀粉青稞利于拓展青稞的应用领域, 从而提高其经济价值。 本研究以114份青藏高原青稞品种(系)为实验材料，通过SDS-PAGE对材料的胚乳淀粉颗粒结合蛋白（SGAPs）进行分离，确定各蛋白的分子量大小、组合类型和多态性等。然后按照国标法测试材料的籽粒总淀粉含量和直链淀粉含量，通过微型糊化粘度仪分析相应的淀粉糊化特性，最后使用显微镜观察比较了青稞的淀粉颗粒形态特征。主要结果如下： 1、114种青稞中共分离出20种不同的SGAP条带，条带分子量为35.00～112.39 KDa，分布频率为12.28～97.37%。材料含有的SGAPs条带数从10到14不等，超过一半的材料含11种SGAP条带。20种条带形成16种组合类型，其中西藏地区青稞包含所有16个组合类型，四川地区青稞包含其中12个组合类型。青藏高原青稞籽粒淀粉颗粒结合蛋白的差异很大，遗传多样性丰富。 2、114份青稞的总淀粉含量、直链淀粉含量、直支淀粉比、峰值粘度、糊化温度和峰值温度的变幅分别为51.26～66.70%、14.64～29.74%、0.17～0.42、194～1135BU、58.8～65.2℃和81.4～92.4℃，相应的平均值分别为59.82%、23.60%、0.31、722.30BU、62.1℃和88.8℃。群体在总淀粉含量、直链淀粉含量、直支淀粉比、峰值粘度、糊化温度和峰值温度上的分布具有明显的正态性；所有胚乳淀粉体的淀粉粒都呈复粒结构。对西藏和四川的材料进行了分组比较， 两地区的青稞在直链淀粉含量和直支淀粉比上的差异达到显著水平。 3、筛选出18份具有特殊淀粉特性的青稞品种，其中5份材料的总淀粉含量超过65%，包括NB63-1、NB67、甘孜白六棱、98221-1和NB63；3份材料的直链淀粉含量大于29%，包括藏青85、藏青3号和喜马拉6号；8份材料的直支淀粉比小于0.25，包括99033-6、春青稞、阿坝330、Jan-03、米麦114、396、NB63-1和92013；7份材料的糊化温度低于60℃，同时材料的峰值粘度大于1000BU,并且峰值温度低于90℃，包括足捉春、Jan-03、阿坝330、米麦114、春青稞、20003和阿青5号。 4、各淀粉特性间存在高度相关性。直链淀粉含量和直支淀粉比与糊化温度成极显著正相关，与峰值粘度成极显著负相关，与A型淀粉粒数量和大小呈负相关。不同SGAPs组合的品种之间，淀粉含量和淀粉糊化特性间差异均达显著水平。SGAP2、SGAP5、SGAP6和SGAP7可能对籽粒直链淀粉含量、直支淀粉比和糊化温度有正向效应；SGAP3、SGAP9∼SGAP20可能对峰值粘度有正向效应。 本研究对青藏高原青稞淀粉资源进行了较为全面的评价，对该区青稞淀粉特性有了系统的认识。研究筛选出的特殊青稞品种可作为青稞育种和青稞淀粉工业应用的潜在资源，淀粉特性差异巨大的众多青稞品种也为拓宽青稞应用领域提供了丰富的资源保障。本研究对部分SGAPs在性质上的鉴定和功能上的初步推断为青稞材料的筛选提供了指导，也为品质育种提供了理论参考。 Hulless barley (naked barley, Hordeum vulgare L.) is a short- season, early maturing crop with a wide range of adaptation. It has been attracting more and more attention due to its superior nutrition and extensive industrial applications. Starch is the main ingredient in hulless barley seeds which makes up 65 percent of hulless barley’s dry weight. The ratio of the amylose/amylopectin and the size, shape, distribution of starch granules can affect the physico-chemical and functional properties of starch, which may turn affect its utilizations. The Qinghai-Tibet Plateau, which is located in southwestern China, is a typical area of vertical agricultural ecosystem and one of the barley origin centers with abundant hulless barley resources. There are little reports about hulless barley in Qinghai-Tibet Plateau at present. To screen and cultivate some characteristic hulless barley can improve its value. An improved SDS-PAGE was used to identify SGAPs combination of 114 hulless barley varieties. Starch content (total starch and amylose starch) was determined according to the standard methods GB5006-85 and GB/T 15683 using PerkinElmer M341 Precision Automatic Polarimeter and UV spectrophotometer 755B respectively. The pasting properties were measured by BRABENDER Micrio Visco-Amylo- Graph 803201. The morphology of starch granules were observed and compared with Axioplan 2 Imaging light microscopy. The following were the results obtained: 1. There were 20 major SGAPs presented in 114 varieties, with the molecular weight ranged from 35.00 to 112.39 KDa, and the frequencies ranged from 12.28% to 97.37%. The number of SGAP bands in each accession varied from 10 to 14, more than half of the population had 11 bands. There were 16 distinct SGAP patterns in the 114 varieties, the Tibet hulless barley had all of the 16 types and the Sichuan hulless barley had 12 types. The results indicated the Qinghai-Tibet Plateau hulless barley had a polymorphism of the SGAPs. 2. The ranges of the total starch content, amylose content, Am/Ap, peak viscosity, pasting temperature and peak temperature of the 114 hulless barley were 51.26～66.70%,14.64～29.74%,0.17～0.42,194～1135BU,58.8～65.2 and 81.4℃～92.4, with an average of ℃59.82%, 23.60%, 0.31, 722.30BU, 62.1 and 88.8,℃℃ respectively. The distributions of the total starch content, amylose content, Am/Ap, peak viscosity, pasting temperature and peak temperature were visibly normal school. All of the amyloplasts in endosperm of varieties showed bimodal size distributions.The main starch properties of hulless barley from Tibet and Sichuan were separated and compared, the differences on amylose content and Am/Ap were obvious. 3. Eighteen accessions which had special starch properties were screened out. Five accessions with total starch content beyond 65%, including NB63-1, NB67, Ganzibailiuleng, 98221-1 and NB63; three accessions, Zangqing85, Zangqing3 and Ximala6, with the highest amylose content (>29%); five accessions with Am/Ap less than 0.25, including 99033-6, Chun Qingke, A Ba 330, Jan-03, Mi Mai114, 396, NB63-1 and 92013; seven accessions had a pasting temperature under 60, ℃meanwhile their peak viscosity beyond 1000BU and their peak temperature under 90℃，including Zu Cuochun, Jan-03, A Ba 330, Mi Mai 114, Chun Qingke, 20003 and A Qing 5. 4. There were high correlations between starch properties. Amylose content and Am/Ap were positively correlated to pasting temperature, negatively correlated to peak viscosity, negatively correlated to the number and granule size of A-type granule. Different SGAP combinations caused significant diversities in starch content and pasting properties. SGAP2, SGAP5, SGAP6 and SGAP7 may have positive effect on amylose content, Am/Ap and pasting temperature; SGAP3, SGAP9∼SGAP20 may have positive effect on peak viscosity. Our research made a comprehensive evaluation on the hulless barley starch from the Qinghai-Tibet Plateau, we can get a systemic understanding. Some special accessions were screened out can be used on the hulless barley breeding lines and industries utilization.The combination of the SGAPs may become a criterion to evaluate the hulless barley endosperm starch quality. Consequently, the results will be good information for further studies on the hulless barley.

Isolation and characterization of a novel variant of HMW glutenin subunit gene from the St genome of Pseudoroegneria stipifolia

The x- and y-type high molecular weight (HMW) glutenin subunits are conserved seed storage proteins in wheat and related species. Here we describe investigations on the HMW glutenin subunits from several Pseudoroegneria accessions. The electrophoretic mobilities of the HMW glutenin subunits from Pd. stipifolia, Pd tauri and Pd strigosa were much faster than those of orthologous wheat subunits, indicating that their protein size may be smaller than that of wheat subunits. The coding sequence of the Glu-1St1 subunit (encoded by the Pseudoroegneria stipifolia accession PI325181) was isolated, and found to represent the native open reading frame (ORF) by in vitro expression. The deduced amino acid sequence of Glu-1St1 matched with that determined from the native subunit by mass spectrometric analysis. The domain organization in Glu-1St1 showed high similarity with that of typical HMW glutenin subunits. However, Glu-1St1 exhibited several distinct characteristics. First, the length of its repetitive domain was substantially smaller than that of conventional subunits, which explains its much faster electrophoretic mobility in SDS-PAGE. Second, although the N-terminal domain of Glu-1St1 resembled that of y-type subunit, its C-terminal domain was more similar to that of x-type subunit. Third, the N- and C-terminat domains of Glu-1St1 shared conserved features with those of barley D-hordein, but the repeat motifs and the organization of its repetitive domain were more similar to those of HMW glutenin subunits than to D-hordein. We conclude that Glu-1St1 is a novel variant of HMW glutenin subunits. The analysis of Glu-1St1 may provide new insight into the evolution of HMW glutenin subunits in Triticeae species. (C) 2007 Elsevier Ltd. All rights reserved.

Morphologically cryptic biological species within the liverwort Frullania asagrayana.

UNLABELLED: PREMISE OF THE STUDY: The Frullania tamarisci complex includes eight Holarctic liverwort species. One of these, F. asagrayana, is distributed broadly throughout eastern North America from Canada to the Gulf Coast. Preliminary genetic data suggested that the species includes two groups of populations. This study was designed to test whether the two groups are reproductively isolated biological species. • METHODS: Eighty-eight samples from across the range of F. asagrayana, plus 73 samples from one population, were genotyped for 13 microsatellite loci. Sequences for two plastid loci and nrITS were obtained from 13 accessions. Genetic data were analyzed using coalescent models and Bayesian inference. • KEY RESULTS: Frullania asagrayana is sequence-invariant at the two plastid loci and ITS2, but two clear groups were resolved by microsatellites. The two groups are largely reproductively isolated, but there is a low level of gene flow from the southern to the northern group. No gene flow was detected in the other direction. A local population was heterogeneous but displayed strong genetic structure. • CONCLUSIONS: The genetic structure of F. asagrayana in eastern North America reflects morphologically cryptic differentiation between reproductively isolated groups of populations, near-panmixis within groups, and clonal propagation at local scales. Reproductive isolation between groups that are invariant at the level of nucleotide sequences shows that caution must be exercised in making taxonomic and evolutionary inferences from reciprocal monophyly (or lack thereof) between putative species.

Salicylic acid biosynthesis is enhanced and contributes to increased biotrophic pathogen resistance in Arabidopsis hybrids.

Heterosis, the phenotypic superiority of a hybrid over its parents, has been demonstrated for many traits in Arabidopsis thaliana, but its effect on defence remains largely unexplored. Here, we show that hybrids between some A. thaliana accessions show increased resistance to the biotrophic bacterial pathogen Pseudomonas syringae pv. tomato (Pst) DC3000. Comparisons of transcriptomes between these hybrids and their parents after inoculation reveal that several key salicylic acid (SA) biosynthesis genes are significantly upregulated in hybrids. Moreover, SA levels are higher in hybrids than in either parent. Increased resistance to Pst DC3000 is significantly compromised in hybrids of pad4 mutants in which the SA biosynthesis pathway is blocked. Finally, increased histone H3 acetylation of key SA biosynthesis genes correlates with their upregulation in infected hybrids. Our data demonstrate that enhanced activation of SA biosynthesis in A. thaliana hybrids may contribute to their increased resistance to a biotrophic bacterial pathogen.

New wild Brassica oleracea L. germplasm at its western distribution limit in the Iberian Peninsula

An exploration and collection mission for wild Brassica oleracea populations was carried out in spring and summer of 2013. The aim of this collection was to expand the number of accessions of wild Brassica oleracea available for basic and applied research in plant breeding. In this paper we report a new accession of wild Brassica oleracea in an unexplored coastal area of Galicia, NW Iberian Peninsula. Details of population ecology and vegetation, soil, climate and geographic data were recorded for this population. The “Endangered” threat category for the region is proposed, and actions for in situ and ex situ conservation are proposed. Seeds will be added to the germplasm collections of University of Santiago de Compostela and Misión Biológica de Galicia (CSIC) for further research on diverse aspects of the dynamics and ecophysiology of the population along with characterization and evaluation of useful traits.

rbcL sequences reveal multiple cryptic introductions of the Japanese red alga Polysiphonia harveyi.

In Europe, the last 20 years have seen a spectacular increase in accidental introductions of marine species, but it has recently been suggested that both the actual number of invaders and their impacts have been seriously underestimated because of the prevalence of sibling species in marine habitats. The red alga Polysiphoniaharveyi is regarded as an alien in the British Isles and Atlantic Europe, having appeared in various locations there during the past 170 years. Similar or conspecific populations are known from Atlantic North America and Japan. To choose between three competing hypotheses concerning the origin of P. harveyi in Europe, we employed rbcL sequence analysis in conjunction with karyological and interbreeding data for samples and isolates of P. harveyi and various congeners from the Pacific and North Atlantic Oceans. All cultured isolates of P. harveyi were completely interfertile, and there was no evidence of polyploidy or aneuploidy. Thus, this biological species is both morphologically and genetically variable: intraspecific rbcL divergences of up to 2.1% are high even for red algae. Seven rbcL haplotypes were identified. The four most divergent haplotypes were observed in Japanese samples from Hokkaido and south-central Honshu, which are linked by hypothetical 'missing' haplotypes that may be located in northern Honshu. These data are consistent with Japan being the centre of diversity and origin for P. harveyi. Two non-Japanese lineages were linked to Hokkaido and Honshu, respectively. A single haplotype was found in all North Atlantic and Mediterranean accessions, except for North Carolina, where the haplotype found was the same as that invading in New Zealand and California. The introduction of P. harveyi into New Zealand has gone unnoticed because P. strictissima is a morphologically indistinguishable native sibling species. The sequence divergence between them is 4–5%, greater than between some morphologically distinct red algal species. Two different types of cryptic invasions of P. harveyi have therefore occurred. In addition to its introduction as a cryptic sibling species in New Zealand, P. harveyi has been introduced at least twice into the North Atlantic from presumed different source populations. These two introductions are genetically and probably also physiologically divergent but completely interfertile.

Tracking biological invasions in space and time: elucidating the invasive history of the green alga Codium fragile using old DNA

With the advent of 'ancient DNA' studies on preserved material of extant and extinct species, museums and herbaria now represent an important although still underutilized resource in molecular ecology. The ability to obtain sequence data from archived specimens can reveal the recent history of cryptic species and introductions. We have analysed extant and herbarium samples of the highly invasive green alga Codium fragile, many over 100 years old, to identify cryptic accessions of the invasive strain known as C. fragile ssp. tomentosoides, which can be identified by a unique haplotype. Molecular characterization of specimens previously identified as native in various regions shows that the invasive tomentosoides strain has been colonizing new habitats across the world for longer than records indicate, in some cases nearly 100 years before it was noticed. It can now be found in the ranges of all the other native haplotypes detected, several of which correspond to recognized subspecies. Within regions in the southern hemisphere there was a greater diversity of haplotypes than in the northern hemisphere, probably as a result of dispersal by the Antarctic Circumpolar Current. The findings of this study highlight the importance of herbaria in preserving contemporaneous records of invasions as they occur, especially when invasive taxa are cryptic.

High-resolution organellar genome analysis of Triticum and Aegilops sheds new light on cytoplasm evolution in wheat

We have utilised polymorphic chloroplast microsatellites to analyse cytoplasmic relationships between accessions in the genera Triticum and Aegilops. Sequencing of PCR products revealed point mutations and insertions/deletions in addition to the standard repeat length expansion/contraction which most likely represent ancient synapomorphies. Phylogenetic analyses revealed three distinct groups of accessions. One of these contained all the non-Aegilops speltoides S-type cytoplasm species, another comprised almost exclusively A, C, D, M, N, T and U cytoplasm-type accessions and the third contained the polyploid Triticum species and all the Ae. speltoides accessions, further confirming that Ae. speltoides or a closely related but now extinct species was the original B-genome donor of cultivated polyploid wheat. Successive decreases in levels of genetic diversity due to domestication were also observed. Finally, we highlight the importance of elucidating longer-term evolutionary processes operating at microsatellite repeat loci.

Genetic variability evaluation in a Moroccan collection of barley, Hordeum vulgare L., by means of storage proteins and RAPDS

The genetic variation existing in a set of barley (Hordeum vulgare L.) landrace samples recently collected in Morocco was estimated. Two kinds of genetic markers, seed storage proteins (hordeins) and random amplified polymorphic DNA (RAPD), were used. Only six out of 31 landraces were subjected to RAPD analysis. Both kinds of markers, RAPD and storage proteins, yielded similar results, showing that the level of variation observed in Moroccan barley was high: all landraces showed variability; 808 different storage protein patterns (multilocus associations) were observed among 1897 individuals (2.32 seeds per association, on average) with an average of 43 multilocus associations per accession. In general, genetic variation within accessions was higher than between accessions. The 100 polymorphic RAPD bands generated by 21 effective primers were able to generate enough patterns to differentiate between uniform cultivars and even between individuals in variable accessions. One of the aims of this work was to compare the effectiveness of RAPD versus storage protein techniques in assessing the variability of genetic resource collections. On average hordeins were more polymorphic than RAPDs: they showed more alternatives per band on gels and a higher percentage of polymorphic bands, although RAPDs supply a higher number of bands. Although RAPD is an easy and standard technique, storage protein analysis is technically easier, cheaper and needs less sophisticated equipment. Thus, when resources are a limiting factor and considering the cost of consumables and work time, seed storage proteins must be the technique of choice for a first estimation of genetic variation in plant genetic resource collections.

Conserved histidine of metal transporter AtNRAMP1 is crucial for optimal plant growth under manganese deficiency at chilling temperatures

Manganese (Mn) is an essential nutrient required for plant growth, in particular in the process of photosynthesis. Plant performance is influenced by various environmental stresses including contrasting temperatures, light or nutrient deficiencies. The molecular responses of plants exposed to such stress factors in combination are largely unknown. 

Screening of 108 Arabidopsis thaliana (Arabidopsis) accessions for reduced photosynthetic performance at chilling temperatures was performed and one accession (Hog) was isolated. Using genetic and molecular approaches, the molecular basis of this particular response to temperature (GxE interaction) was identified. 

Hog showed an induction of a severe leaf chlorosis and impaired growth after transfer to lower temperatures. We demonstrated that this response was dependent on the nutrient content of the soil. Genetic mapping and complementation identified NRAMP1 as the causal gene. Chlorotic phenotype was associated with a histidine to tyrosine (H239Y) substitution in the allele of Hog NRAMP1. This led to lethality when Hog seedlings were directly grown at 4 degrees C. 

Chemical complementation and hydroponic culture experiments showed that Mn deficiency was the major cause of this GxE interaction. For the first time, the NRAMP-specific highly conserved histidine was shown to be crucial for plant performance.

Genome Wide Association Mapping of Grain Arsenic, Copper, Molybdenum and Zinc in Rice (Oryza sativa L.) Grown at Four International Field Sites

The mineral concentrations in cereals are important for human health, especially for individuals who consume a cereal subsistence diet. A number of elements, such as zinc, are required within the diet, while some elements are toxic to humans, for example arsenic. In this study we carry out genome-wide association (GWA) mapping of grain concentrations of arsenic, copper, molybdenum and zinc in brown rice using an established rice diversity panel of,300 accessions and 36.9 k single nucleotide polymorphisms (SNPs). The study was performed across five environments: one field site in Bangladesh, one in China and two in the US, with one of the US sites repeated over two years. GWA mapping on the whole dataset and on separate subpopulations of rice revealed a large number of loci significantly associated with variation in grain arsenic, copper, molybdenum and zinc. Seventeen of these loci were detected in data obtained from grain cultivated in more than one field location, and six co-localise with previously identified quantitative trait loci. Additionally, a number of candidate genes for the uptake or transport of these elements were located near significantly associated SNPs (within 200 kb, the estimated global linkage disequilibrium previously employed in this rice panel). This analysis highlights a number of genomic regions and candidate genes for further analysis as well as the challenges faced when mapping environmentally-variable traits in a highly genetically structured diversity panel.

Mechanisms of plant resistance to metal and metalloid ions and potential biotechnological applications

Metal and metalloid resistances in plant species and genotypes/accessions are becoming increasingly better understood at the molecular and physiological level. Much of the recent focus into metal resistances has been on hyperaccumulators as these are excellent systems to study resistances due to their very abnormal metal(loid) physiology and because of their biotechnological potential. Advances into the mechanistic basis of metal(loid) resistances have been made through the investigation of metal(loid) transporters, the construction of mutants with altered metal(loid) transport and metabolism, a better understanding of the genetic basis of resistance and hyperaccumulation and investigations into the role of metal(loid) ion chelators. This review highlights these recent advances. © Springer 2005.

Analysis of genetic diversity in Portuguese Ceratonia siliqua L. cultivars using RAPD and AFLP markers

Although carob (Ceratonia siliqua L.) is of great economic importance little is still known about the pattern of genetic variation within this species. Morphological characteristics based on 31 fruit and seeds of continuous characters determinant for agro-industrial uses, were compared with RAPD and AFLP markers for assessing genetic distances in 68 accessions of carob trees, from different cultivars, varieties and eco-geographic regions of Algarve. Eighteen selected RAPD primers applied to the 68 accessions produced a total of 235 fragments ranging from 200 to 2000 bp, of which 93 (40%) were polymorphic. Four AFLP selective primer combinations generated a total of 346 amplification fragments of which 110 were polymorphic. The average level of polymorphism based on four primer combinations was 31.8%. The phenetic trees based on RAPD and AFLP analyses gave high co-phenetic correlation values, and were found to be consistent in general with the analysis of morphological data, carried out on the same accessions. A number of RAPD and AFLP markers were found to be diagnostic for ‘Canela’ cultivar and 13 wild ungrafted trees.

Estudo morfológico e filogenético das subespécies Daucus carota ssp. Azoricus e Daucus carota ssp. Maritimus na ilha de S. Miguel

Dissertação de Mestrado, Biodiversidade e Biotecnologia Vegetal, 17 de Março de 2015, Universidade dos Açores.

Ethnobotanique de la Nation crie d'Eeyou Istchee et variation géographique des plantes médicinales antidiabétiques

Le diabète de type 2 affecte en moyenne 29% de la population adulte crie d’Eeyou Istchee (CEI). Afin d’identifier les plantes médicinales possédant un potentiel antidiabétique, des interviews ont été réalisés dans les communautés CEI de Wemindji et Oujé-Bougoumou. Utilisant une approche quantitative, les espèces mentionnées ont été classées et comparées à la pharmacopée des communautés avoisinantes. Seize et 25 plantes ont été mentionné à Wemindji et Oujé-Bougoumou, respectivement. Sept nouvelles espèces de plantes et une de champignon se sont ajoutées à la liste des espèces à potentiel antidiabétique, bien que la plupart de celles mentionnées pendant les interviews soit en communes à la pharmacopée CEI générale, démontrant ainsi leur importance culturelle. Des analyses phytochimiques sur deux de ces espèces, Rhododendron groenlandicum et Sarracenia purpurea, ont été réalisées à partir d’échantillons récoltés à différents endroits du territoire eeyouch. Bien qu’aucun patron n’ait été détecté dans la variation des composantes biologiquement actives chez S. purpurea, les composés phénoliques chez R. groenlandicum, particulièrement la (+)-catéchine, l’(-)-epicatéchine et la quercétine-3-galactoside, varient spatialement en fonction de paramètres d’insolation telles la radiation solaire ou la photopériode. Les échantillons de cette dernière espèce, testés in vitro dans le bioessai de l’adipogenèse des cellules adipocytes murines 3T3-L1, augmentent l’accumulation intracellulaire des triglycérides, leur conférant ainsi une activité diabétique semblable à la rosiglitazone. Cependant, cette activité était plus faible dans les échantillons à haute teneur en quercétine, cela pouvant ainsi avoir un impact sur la qualité d'un produit de santé naturel fabriqué à partir de cette espèce.