Short catkin1, a novel mutant of Castanea mollissima, is associated with programmed cell death during chestnut staminate flower differentiation

The fact that male flowers far outnumber female flowers is a factor that limits nut yield in Chinese chestnut. A naturally occurring mutation of male catkins was found on a single branch of a Chinese chestnut tree in the mountains near Beijing, China. The mutation was named short catkin1 (sck1). The catkin length of sck1 was only 1/6 to 1/8 that of the wild-type male catkins on the same tree. The mutation was associated with a greater number of female flowers and increased yield. Observations on the development of male catkins with the sck1 phenotype showed that the distal part of the catkins aborted at the stage of chestnut staminate flower differentiation. Further research using transmission electron microscope analysis showed that the process of cell death in sck1 catkins had the typical characteristics of programmed cell death at the subcellular level, such as condensed chromatin, dissolved nucleolus, degraded karyoplasm, burst karyotheca, and disintegrated chloroplasts or mitochondria. Significantly, DNA laddering was detected in tissues of sck1 catkins. In conclusion, the results showed that sck1 was associated with PCD.     

短雄花序板栗芽变的AFLP分析

 利用AFLP分子标记技术对板栗短雄花序芽变及其母株进行了初步研究, 72种引物组合共产生5 129条清晰谱带, 多态性位点53个, 多态性2.05% , 相似性系数0.9897, 差异片段主要集中于300～900 bp之间。研究表明短雄花序芽变和对照个体之间变异与遗传物质的改变相关, 其中一些可能与雄花序变短的基因有关。     

Effects of ethephon,GA3 and nutrient elements on sex expression of Chinese chestnut

The effects of ethephon or GA₃ on sex expression of Chinese chestnut (Castanea mollissima BL.) were opposite to the effects of these plant growth regulators on cucumber, pumpkin and Chinese white-flowered gourd. Ethephon at 100–500 mg 1^?1 inhibited the development of female flowers and promoted that of male flowers. GA₃ at 50–500 mg 1^?1 caused a significantly lower number of male inflorescences, but increased the number of female flowers. Urea applications decreased the number of male inflorescences and increased the number of female flowers. For plants grown on phosphorous-deficient soils, fertilizing with superphosphate also increased the number of female flowers. Spraying trees with GA₃, urea or a mixture of both increased the number of female flowers. Therefore, it is proposed that such applications should be employed at the onset of differentiation of flower primordia of Chinese chestnut to increase the ratio of female to male flowers.     

Novel genomic and EST-derived SSR markers in Japanese chestnuts

A total of 366 novel simple sequence repeat (SSR) markers were developed in Japanese chestnut (Castanea crenata), comprising 220 genomic SSRs derived from enriched genomic libraries and 146 expressed sequence tag (EST)–SSRs obtained from large-scale EST sequencing analysis. Thirty accessions, comprising Japanese, Chinese (C. mollissima), European (Castanea sativa), and American chestnuts (Castanea dentata), were used for evaluation of SSR polymorphism and transferability across species. The EST–SSRs showed less polymorphism than the genomic SSRs and were more transferable. The mean observed heterozygosity (H_O) and the mean expected heterozygosity (H_E) of genomic SSRs in the Japanese chestnuts were 0.63 and 0.68, respectively; those of EST-SSRs were each 0.47. Although about 80% of the genomic SSRs were amplified in all 4 species, more than 95% of the EST–SSRs were transferable across all 4 species. The many novel SSRs developed in this study will be applicable for the construction of genetic linkage maps, QTL analysis of phenotypic traits, high-throughput genotyping of marker-assisted selection, and association genetics.     

Rooting stem cuttings of Chinese chestnut

Rooting of Chinese chestnut (Castanea mollissima Blume.) stem cuttings following treatment with indole-3-butyric acid (IBA) was investigated. Tip and basal cutting from vigorous epicormic shoots and terminal shoots with 2–3 cm of the previous year's wood were taken from mature regions of trees approximately 40 years of age. Cuttings were dipped for 5 s in an aqueous solution of either 3 or 6 g l^?1 IBA. Rooting occurred only in the basal softwood cuttings. Average rooting of 33.5, 5.0 and 1.6% for ‘AU-Leader’, ‘AU-Homestead’ and ‘AU-Cropper’, respectively, was obtained using the 3 g l^?1 IBA solution, and 35.0, 6.7 and 3.3%, respectively, using the 6 g l^?1 IBA solution.     

罗田甜柿Ty1-copia类逆转座子RNaseH-LTR序列的分离和特性分析

逆转座子序列信息的获得,对了解其在基因组中的行为及系统学研究有重要价值。本试验从罗田甜柿（Diospyros kaki Thunb. 'Luotian-tianshi'）基因组中分离31个RNaseH-LTR（Long Terminal Repeat,长末端重复）序列,并利用IRAP（Inter-Retrotransposon Amplified Polymorphism,逆转座子间扩增多态性）技术对部分序列相应的逆转座子家族在柿属植物中的转座活性及分布情况进行初步探讨。序列分析结果表明,至少有10个Ty1-copia类逆转座子家族得到扩增;其家族间普遍表现高度异质,碱基替换、插入或缺失突变,以及翻译成氨基酸后发生不同程度的终止密码子突变、氨基酸取代和移框突变等,是产生高异质性的原因;此外,其家族内部某些序列间的相似性极高,可能是寄主基因组与逆转座子间互惠关系的体现。应用部分逆转座子引物的IRAP分析结果表明,相应的逆转座子家族在柿属植物中普遍存在,其分布广泛,拷贝数高,转座活性强,具有进一步开发为多种逆转座子分子标记的潜力。     

枇杷LTR类反转录转座子RT序列的克隆及分析

[目的]揭示枇杷LTR类反转录转子座RT序列在枇杷基因组中的异质性,为枇杷转座子进化和多样性研究提供依据.[方法]以普通枇杷野生种质的叶片为材料,通过简并引物从枇杷基因组中扩增得到Ty1-copia和Ty3-gypsy类反转录转座子反转录酶序列,并对其序列变化特点进行生物信息学分析.[结果]最终获得38条Ty1-cop...     

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.     

应用抑制性消减杂交分离板栗短雄花序芽变相关基因

利用抑制性消减杂交技术分离了板栗正常花序与芽变花序之间表达的差异cDNA片段。以芽变板栗雄花序cDNA为试验方(tester),以对照板栗雄花序cDNA为驱动方(driver),构建了一个板栗芽变花序消减文库。菌落PCR显示插入片段主要在100～1000 bp,文库质量良好。通过对文库阳性克隆随机测序共得到了30个有效EST序列。利用Blast同源性比较,其中一些EST可能与板栗雄花序发育中变短的生物学形态形成密切相关,为后续板栗短雄花序芽变相关基因的研究奠定了基础。     

Presence and abundance of the Eurasian nuthatch <Emphasis Type="Italic">Sitta europaea</Emphasis> in relation to the size,isolation and the intensity of management of chestnut woodlands in the NW Iberian Peninsula

Throughout most of the north-west Iberian Peninsula, chestnut (Castanea sativa) woods are the principal deciduous woodland, reflecting historical and ongoing exploitation of indigenous forests. These are traditionally managed woodlands with a patchy distribution. Eurasian nuthatches (Sitta europaea) inhabit mature deciduous woods, show high site fidelity, and are almost exclusively found in chestnut woods in the study area. We studied the presence and abundance of nuthatch breeding pairs over two consecutive years, in relation to the size, degree of isolation and intensity of management of 25 chestnut woods in NW Spain. Degree of isolation was assessed in view of the presence of other woodland within a 1-km band surrounding the study wood. Wood size was the only variable that significantly predicted the presence of breeding pairs (in at least one year, R ² = 0.69; in both years, R ² = 0.50). The number of pairs was strongly predicted by wood size, isolation and management (R ² = 0.70 in 2004; R ² = 0.84 in 2005); interestingly, more isolated woods had more breeding pairs. Breeding density was likewise significantly or near-significantly (P ≤ 0.1) higher in small isolated woods, which is possibly attributable to lower juvenile dispersal in lightly forested areas and/or to lower predator density in smaller and more isolated patches. Breeding density was higher (though not significantly so) in more heavily managed woods, possibly due to the presence of larger chestnut crops and larger trees (with higher nuthatch prey abundance). Our findings highlight the complexity of the relationships between the patch properties and the three studied levels (presence, number and density of pairs), and also the importance of traditionally managed woodlands for the conservation of forest birds.     

Morphology and anatomy of pollen grains from male-fertile and male-sterile cultivars of chestnut (Castanea sativa Mill.)

Summary

Pollen morphology and ultrastructure are described for four male-fertile cultivars (‘Firdola’, ‘Karamehmet’, ‘Sar?a?lama’, and ‘Hac?ömer’) and two male-sterile cultivars (‘Osmanog?lu’ and ‘Vakit Kestanesi’) of chestnut (Castanea sativa Mill.) using light microscopy and both scanning and transmission electron microscopy. Pollen grains of both male-fertile and male-sterile chestnut cultivars are tricolporate, the germinal furrow extending almost the full length of the grain axis. Pollen grains have a slightly reticulate exine. Pollen grain length varied from 13.33 – 21.30 µm, and decreased significantly in the male-sterile cultivars. Three different pollen shapes were observed among the cultivars: prolate, perprolate, and sub-prolate. The ultrastructure of the pollen grains did not differ between male-fertile and male-sterile cultivars. Intine, exine and total wall thickness (exine + intine) of pollen grains were determined as: 83.2 – 153.1 nm, 432.8 – 520.0 nm, and 516.0 – 651.6 nm, respectively; and variations were significant (P ≤ 0.05) among cultivars. The percentages of in vitro germination of pollen grains of male-fertile cultivars were between 11 – 78%, and the variations among cultivars were significant (P ≤ 0.05). The percentages of empty pollen grains observed among cultivars ranged from 3 – 32%. The correlation coefficient between the percentage of normal pollen and the germination rate was r = 0.898 (P ≤ 0.01).     

甜瓜属REMAP分子标记体系的建立及应用

基于甜瓜属Ty1-copia类逆转座子的长末端重复序列信息,结合ISSR引物序列,在体系优化的基础上建立了适用于甜瓜属物种的REMAP(retrotransposon-microsatellite amplified polymorphism)标记体系。对46份甜瓜属不同类型材料进行聚类分析,证明该标记体系能有效检测甜瓜属不同类型材料间的多态性,可以用于甜瓜属作物品种鉴定及指纹图谱构建。     

Chestnut (Castanea sativa Mill.) genotype identification: An artificial neural network approach

Summary

The potential use of the artificial neural networks (ANNs) for characterization and identification of seventeen chestnut (Castanea sativa Mill.) accessions, belonging to the ``marrone''-type and ``chestnut''-type, was investigated in genotypes originating from regions of Italy. Different back-propagation neural networks (BPNN) were built on the basis of image analysis parameters of the leaves, for two tasks of chestnut classification. In the first case a BPNN was built and trained to differentiate the 17 accessions of chestnut. In the second case a BPNN was conceived to distinguish between the ``marrone'' and ``chestnut'' types. BPNN produced a clear identification of all the accessions except in the case of `Garrone nero', `Garrone rosso' and `Tempuriva', which showed almost the same output diagram. Cluster analysis separated the 17 chestnut genotypes into four main groups whose differences were related to the original sources of the genotypes and to the type of affiliation (``marrone''-type or ``chestnut''-type). Artificial neural network technique was also able to discriminate between ``marrone''-type and ``chestnut''-type accessions. Qualitative and quantitative rules for the image analysis parameters, useful for classifying chestnut accessions into these two types, were obtained. On the whole the relative importance of the leaf parameters reveals that ``typical'' leaves for ``marrone''-type are more elongated, of a darker colour and with a higher perimeter/area ratio than the leaves of the ``chestnut''-type.     

Woody species composition of chestnut stands in the Northern Apennines: the result of 200 years of changes in land use

Chestnut stands (orchards and coppices) are among the most typical elements of the southern European mountain landscape and a protected habitat (9260 Castanea sativa woods) according to the European Union (Directive 92/43/EEC). As an anthropogenic landscape, they require specific measures to address preservation or to guide their evolutionary trend. In the Northern Apennines, a landscape multiscalar-multitemporal approach was adopted to highlight factors that have acted on the evolution of this habitat and which still might affect either its preservation or its evolutionary dynamics. Using a diachronic GIS-approach, we analyzed old cadastral maps (drawn up 200 years ago), and aerial photographs. Both the present distribution pattern of the woody species and the incidence of important chestnut diseases were also surveyed. The factors explaining the current extent and species composition of the local chestnut forests confirm their status as an anthropogenic habitat. The present landscape distribution of chestnut woods is heavily linked to past human settlements. Chestnut blight and ink disease are more an indirect reason for past felling activities than an actual direct cause of damage to trees, because of the hypovirulence spread and the limited incidence of the ink disease. Vegetation dynamics of abandoned chestnut forests evolved only partly towards deciduous Beech and Hop Hornbeam stands, thus suggesting both the possibility of a recovery of this cultivation and the need for new criteria for its management.     

牡丹Ty3-gypsy 类反转录转座子反转录酶序列的克隆及分析

 根据Ty3-gypsy反转录转座子反转录酶的保守序列设计简并引物,从中原牡丹（Paeonia suffruticosa Andrews）品种‘洛阳红’和野生种卵叶牡丹（Paeonia qiui Y. L. Pei et D. Y. Hong）中扩增出430 bp左右的目标片段。目的条带经回收、克隆、测序及相关生物信息学软件进行序列分析后,获得了13条来自牡丹的Ty3-gypsy反转录转座子反转录酶序列。这些核苷酸序列具有较高的异质性,主要表现为缺失突变,序列长度变化范围为412 ~ 446 bp,同源性范围为71.5% ~ 94.8%。翻译成氨基酸后,有12条序列出现1 ~ 9个不同程度的终止密码子突变,3条序列出现移框突变。其核苷酸序列经过系统聚类后可分为6个家族。将其氨基酸序列与已登录的不同物种Ty3-gypsy反转录转座子反转录酶的氨基酸序列进行聚类分析,结果表明与其他植物具有较高的同源性,表明它们间可能存在着Ty3-gypsy反转录转座子的横向传递。     

Identification and structure of six members of the hexokinase gene family in Vitis vinifera: Cloning,expression, and functional analysis of a putative chloroplast stromal-type hexokinase

Summary

Hexokinase catalyses the first step in the metabolism of glucose, but has also been proposed to be involved in sugar sensing and signalling in both yeasts and plants. The substrates of hexokinase such as glucose (Glc) and fructose (Fruc) are also the most important sugars during grape berry ripening. The grapevine proteome database was analysed to investigate the roles of hexokinases in grape berry growth and development. Six hexokinase genes displaying high nucleotide sequence identity (72 – 87%) with hexokinase genes in other species were identified. Most of the Vitis vinifera hexokinase (VvHXK) genes had a highly conserved genomic structure consisting of nine exons and eight introns. A search for cis–regulatory elements in the promoter regions of all six hexokinase genes revealed that most were probably regulated by light, sugar, phytohormones, or abiotic stress. The isolation and expression of a hexokinase cDNA from V. vinifera ‘Sultanine’, named VvHXK3, was also reported. Analysis of the predicted amino acid sequence of VvHXK3 suggested that the protein could be a chloroplast-stromal hexokinase with a possible transit peptide cleavage site after amino acid residue 26. The VvHXK3 gene was differentially expressed in a variety of organs including berries, leaves, roots, and pollen, but its expression was highest in berries during their early development and at the start of ripening. To determine its function, VvHXK3 cDNA was expressed in a triple mutant yeast strain that lacked the ability to phosphorylate Glc and Fru and, therefore, was unable to grow on these sugars as sole carbon source. Mutant yeast cells that expressed VvHXK3 grew on both Glc and Fru, indicating that VvHXK3 could complement this mutant and had hexokinase activity.     

Cloning,characterisation, and expression analysis of an oleosin gene in coconut (Cocos nucifera L.) pulp

Summary

Oleosins are structural proteins found in oil bodies, organelles found in the cells of plant tissues with a high oil content that undergo extreme desiccation as part of their maturation process. Oleosins stabilise oil bodies. In this paper, a full-length cDNA sequence homologous to oleosin, a seed-storage oil-body protein, from coconut (Cocos nucifera L.) was identified from cDNA libraries during fruit development and characterised. The gene, termed Coco-Ole, contained an open reading frame of 375 bp encoding a polypeptide of 125 amino acids. The predicted amino acid sequence of coconut oleosin had a molecular mass of 13.0 kDa, and showed 92% (AAF76238.1) and 67% (AAC02239.1) sequence similarity to oil palm (Elaeis guineensis Jacq.) and rice (Oryza sativa) oleosin proteins, respectively. The amino acid sequence clustered in the same branch as oil palm in the cladogram, but was distant from other species. The results of semi-quantitative RT-PCR indicated that the Coco-Ole gene was expressed only in the pulp, and its expression increased significantly during pulp development. Compared with fluctuations in oil content, expression of the Coco-Ole gene was consistent with the anabolism of oil during pulp development. The cloning and sequencing of the Coco-ole gene provides a new marker for studies on oil body biogenesis and fruit development in coconut.     

板栗新品种‘燕龙’

‘燕龙’是从实生燕山板栗中选育出的新品种,经过无性系后代遗传性测定、区试及示范,表现出寡雄、高产、个大、色好、质优、可短截、适于密植等特性。