Interaction of endodormancy and paradormancy in raspberry (Rubus idaeus L.)

Bud break in protected Northern European raspberry crops is often poor and uneven with many of the sub-apical buds remaining in a dormant state. In order to improve bud break and therefore yields, the mechanism controlling bud dormancy must be determined. Canes of the biennial cultivar ‘Glen Moy’ were forced as isolated single nodes, trisections, or as intact canes after different lengths of cold storage chill unit (CU) accumulation in order to determine whether the buds were in an endodormant or paradormant state. The results showed that buds on the lower parts of the intact canes remained in a dormant state long after buds from higher up the intact cane and also the single nodes from all parts of the cane had emerged from the deepest phase of endodormancy. This would imply that these buds were being held in a paradormant state until large amounts of chilling units (>1000 CU) had been accumulated. The trisected cane portions revealed almost no significant differences in bud break levels throughout the experiment when compared with the single nodes. This suggests that removal of the apical part of the cane would be effective in improving bud break by reducing the paradormant condition. A period of secondary dormancy was also observed in the intact canes which may also exacerbate the poor bud break observed in protected crops. This was not seen in the single nodes or the trisected canes which indicates that treatments which reduce paradormancy may also minimise the risk of secondary dormancy. By identifying the phase of bud dormancy which causes poor bud break, attention can now be focused on methods which overcome paradormancy in protected crops. Such methods might include tipping (removal of the cane apex), horizontal training methods, more efficient chilling methods, and chemical treatments.     

Co-ordinated gene expression during phases of dormancy release in raspberry (Rubus idaeus L.) buds

Bud break in raspberry (Rubus idaeus L.) is often poor and uneven, with many of the subapical buds remaining in a dormant state. In order to determine the dormancy status of raspberry buds, an empirical measure of bud burst in a growth-permissive environment following exposure to chilling (4 degrees C cold storage) was developed. For cv. Glen Ample, percentage bud burst in intact canes and isolated nodes was recorded after 14 d. Isolated nodes (a measure of endodormancy) achieved 100% bud burst after approximately 1500 h chilling whereas buds on intact plants (combined endo- and paradormancy) required an additional 1000 h chilling. A microarray approach was used to follow changes in gene expression that occurred during dormancy transition. The probes for the microarrays were obtained from endodormant and paradormant raspberry bud cDNA libraries. The expression profiles of 5300 clones from these libraries were subjected to principal component analysis to determine the most significant expression patterns. Sequence analysis of these clones, in many cases, enabled their functional categorization and the development of hypotheses concerning the mechanisms of bud dormancy release. Thus a set of novel candidates for key dormancy-related genes from raspberry buds have been identified. Bud dormancy is fundamental to the study of plant developmental processes and, in addition, its regulation is of significant economic importance to fruit and horticultural industries.     

Molecular and biochemical characterization of benzalacetone synthase and chalcone synthase genes and their proteins from raspberry (Rubus idaeus L.)

Two new members of the polyketide synthase (PKS) gene family (RiPKS4 and RiPKS5) were cloned from raspberry fruits (Rubus idaeus L., cv Royalty) and expressed in Escherichia coli. Characterization of the recombinant enzyme products indicated that RiPKS4 is a bifunctional polyketide synthase producing both 4-hydroxybenzalacetone and naringenin chalcone. The recombinant RiPKS4 protein, like the native protein from raspberry fruits [W. Borejsza-Wysocki, G. Hrazdina, Plant Physiol. 1996;110: 791-799] accepted p-coumaryl-CoA and ferulyl-CoA as starter substrates and catalyzed the formation of both naringenin chalcone, 4-hydroxy-benzalacetone and 3-methoxy-4-hydroxy-benzalacetone. Although activity of RiPKS4 was higher with ferulyl-CoA than with p-coumaryl-CoA, the corresponding product, 3-methoxy-4-hydroxy phenylbutanone could not be detected in raspberries to date. Sequence analysis of the genes and proteins suggested that this feature of RiPKS4 was created by variation in the C-terminus due to DNA recombination at the 3′ region of its coding sequence. RiPKS5 is a typical chalcone synthase (CHS) that uses p-coumaryl-CoA only as starter substrate and produces naringenin chalcone exclusively as the reaction product.     

The isolation of RNA from raspberry (Rubus idaeus) fruit

Previous attempts to extract high-quality, total RNA from raspberry (Rubus idaeus) fruits using published protocols have proven to be unsuccessful. Even the use of protocols developed for the extraction of RNA from other fruit tissue has resulted in low yields (1) or the isolation of degraded RNA (2). Here, we report on the development of a quick and simple method of extracting total RNA from raspberry fruit. Using this method, high yields of good quality, undegraded RNA were obtained from fruit at all stages of ripening. The RNA is of sufficient quality for northern analysis and cDNA library construction.     

Identification of quantitative trait loci for cane splitting in red raspberry (Rubus idaeus)

Cane splitting, a normal feature of raspberry growth, can lead to plant infestation by cane midge followed by fungal infection, with losses in yield of up to 50 % if left untreated. The extent of splitting in the Latham × Glen Moy reference mapping population was assessed over six years and in three environments and quantitative trait loci (QTL) were identified across linkage groups (LG) 2, 3, 5 and 6. Cane splitting QTL on LG 3 and 5 co-locate with QTL for plant vigour. The cane splitting QTL on LG 6 is associated with the QTL for resistance to root rot caused by Phytophthora rubi. Broad-sense heritability for cane splitting ranged from 25.6 % in 2007 to 49.1 % in 2008 in this population. Season and environment were also found to influence cane splitting in this population. Several genes involved in general plant growth and in defence responses lie within these QTL. This is a first step towards identifying the genetic basis of cane splitting in raspberry and the development of genetic markers for use in raspberry breeding programmes.     

The construction of a genetic linkage map of red raspberry (Rubus idaeus subsp. idaeus) based on AFLPs, genomic-SSR and EST-SSR markers

Breeding in raspberry is time-consuming due to the highly heterozygous nature of this perennial fruit crop, coupled with relatively long periods of juvenility. The speed and precision of raspberry breeding can be improved by genetic linkage maps, thus facilitating the development of diagnostic markers for polygenic traits and the identification of genes controlling complex phenotypes. A genetic linkage map (789 cM) of the red raspberry Rubus idaeus has been constructed from a cross between two phenotypically different cultivars; the recent European cultivar Glen Moy and the older North American cultivar Latham. SSR markers were developed from both genomic and cDNA libraries from Glen Moy. These SSRs, together with AFLP markers, were utilised to create a linkage map. In order to test the utility of the genetic linkage map for QTL analysis, morphological data based on easily scoreable phenotypic traits were collected. The segregation of cane spininess, and the root sucker traits of density and spread from the mother plant, was quantified in two different environments. These traits were analysed for significant linkages to mapped markers using MapQTL and were found to be located on linkage group 2 for spines and group 8 for density and diameter. The availability of co-dominant markers allowed heterozygosities to be calculated for both cultivars.     

Vigour control, an integrated approach to cane, pest and disease management in red raspberry (Rubus idaeus)

The first flush of young canes of red raspberry (Rubus idaeus) was removed at different dates in spring, using the contact herbicide dinoseb. In comparison with untreated plots, cane removal increased fruit yield, controlled excessive cane vigour, and improved the health status of vegetative canes in a plantation infested by raspberry cane midge (Resseliella theobaldi). At the end of the growing season vegetative canes on treated plots were shorter and thinner, and sustained less physical injury than those on untreated plots. Access to the fruit at harvest was also improved. Reduced competition between fruiting and vegetative canes increased yield in the year of treatment by an average of 35%. Yield was not affected by date of cane removal, but growth of replacement cane was reduced below optimum when first flush cane was removed after mid-May. The later the canes were removed, the less was the incidence of pest and diseases. Cankers and lobate vascular lesions (‘patches’) resulting, respectively, from the feeding of first and second generation midge larvae (with associated fungi) affected fewer canes in treated than in untreated plots. Significantly fewer live larvae of R. theobaldi were recovered in the following winter from soil in plots treated on or after 11 May than from untreated plots. The main effect of vigour control on R. theobaldi and midge blight was that replacement canes provided fewer egg-laying sites (natural splits) than did the first flush canes on untreated plots. The incidence of spreading vascular lesions (‘stripes’) attributed to Leptosphaeria coniothyrium infecting either physical wounds (cane blight) or midge feeding wounds (midge blight) was substantially less in treated than untreated plots. Cane botrytis (Botrytis cinerea) and spur blight (Didymella applanata) were also less common in treated plots. Interactions between vigour control and pest and disease incidence are discussed in relation to the efficient management of cv. Glen Clova in eastern Scotland.     

Effects of reflective groundcovers on ground beetles (Coleoptera: Carabidae) in red raspberry (Rubus idaeus) cropping systems

Reflective groundcovers are management tools used in temperate regions where light can be a limiting factor to crop productivity. Although their effects on crops have been extensively studied, little is known about their ecological impacts on ground‐dwelling organisms. Theoretically, reflective groundcovers add structural complexity to the system and thus have the potential to create refuges for ground‐dwelling invertebrates. At the same time, groundcovers can create abiotic conditions underneath them that could potentially cause declines in the abundance and richness of ground‐dwelling invertebrates. During the summers of 2006 and 2007, white, woven, reflective polymer groundcovers were placed in two red raspberry (Rubus idaeus L.) cropping systems (organic and conventional) in south‐eastern New Brunswick to assess their effects on ground beetle richness, activity density and overall assemblage structure. A total of 8723 ground beetles belonging to 23 species were collected using pitfall traps. In contrast with the combined effects of cropping systems and sampling years, the use of a groundcover accounted for a small proportion of the variance in trap catches and caused little change in ground beetle richness, activity density and overall assemblage structure. Based on these results, we suggest that reflective groundcovers constitute promising management tools to increase light environment quality in row crops without being detrimental to ground beetle species that may contribute to pest suppression.     

The Histology of Seed Development in Diploid and Tetraploid Raspberries (Rubus idaeus L.)

Ovules of the raspberry, Rubus idaeus L., resembled those ofother Rubus species in being unitegmic, crassinucellate, andanatropous. In one cultivar, ‘M. 69/139’, both eggcell and embryo were inserted slightly laterally to the micropyle.Of the tetraploid forms, 4n M. 69/139 showed a proportion ofembryo sacs with delayed differentiation, and 4n ‘M. Jewel’some with four antipodal cells. Embryo development showed littleuniformity in timing, but followed the Asterad type. Occasional delayed endosperm development seemed to be a varietalpeculiarity of M. 69/139. At maturity, the ergastic materialin healthy endosperms consisted of oil globules and aleurongrains. The hypostase formed a projecting pedestal below theendosperm at 8 days. The inner epidermal layer of the testaappeared specialized, both in the early stages of seed growth,when it is suggested that it functioned as an endodermis, andat maturity, when it appeared to form an osmotic barrier.     

Construction of black (Rubus occidentalis) and red (R. idaeus) raspberry linkage maps and their comparison to the genomes of strawberry, apple, and peach

The genus Rubus belongs to the Rosaceae and is comprised of 600-800 species distributed world-wide. To date, genetic maps of the genus consist largely of non-transferable markers such as amplified fragment length polymorphisms. An F(1) population developed from a cross between an advanced breeding selection of Rubus occidentalis (96395S1) and R. idaeus 'Latham' was used to construct a new genetic map consisting of DNA sequence-based markers. The genetic linkage maps presented here are constructed of 131 markers on at least one of the two parental maps. The majority of the markers are orthologous, including 14 Rosaceae conserved orthologous set markers, and 60 new gene-based markers developed for raspberry. Thirty-four published raspberry simple sequence repeat markers were used to align the new maps to published raspberry maps. The 96395S1 genetic map consists of six linkage groups (LG) and covers 309 cM with an average of 10 cM between markers; the 'Latham' genetic map consists of seven LG and covers 561 cM with an average of 5 cM between markers. We used BLAST analysis to align the orthologous sequences used to design primer pairs for Rubus genetic mapping with the genome sequences of Fragaria vesca 'Hawaii 4', Malus × domestica 'Golden Delicious', and Prunus 'Lovell'. The alignment of the orthologous markers designed here suggests that the genomes of Rubus and Fragaria have a high degree of synteny and that synteny decreases with phylogenetic distance. Our results give unprecedented insights into the genome evolution of raspberry from the putative ancestral genome of the single ancestor common to Rosaceae.     

Chemical composition of the epicuticular wax from the fruits of Eucalyptus globulus

The chemical composition of the epicuticular wax from the fruits of Eucalyptus globulus was studied by GC-MS before and after alkaline hydrolysis. The wax had two main components, ursolic acid and tritriacontan-16,18-dione, together with several other triterpenic acids. After alkaline hydrolysis, a large increase in the amounts of triterpenic acids and fatty acids (particularly in hexadecanoic acid) was observed, suggesting that these components were present predominantly in esterified forms in the fruit wax. Six compounds were isolated from the fruits by preparative chromatography, and were identified as 8-desmethyleucalyptin, sesamin, tritriacontan-16,18-dione, ursolic acid, 3beta-hydroxyurs-11-en-13beta(28)-olide (ursolic acid lactone) and 3beta,11alpha-dihydroxyurs-12-en-28-oic acid, the latter of which was identified for the first time.     

Establishment of callus and cell suspension cultures of raspberry (Rubus idaeus cv. Royalty)

Callus cultures were initiated from leaf sections of raspberry (Rubus idaeus L.) cv. Royalty. Explants from younger leaves produced significantly more calli than those from older leaves. Anderson's salt mixture was more efficient for callus induction than the Murashige-Skoog medium. The best propagation and growth of calli was observed on Anderson's medium supplemented with 9 M 2,4-dichlorophenoxyacetic acid, 4.9 M indolebutyric acid and 4.9 M 6-(dimethylallylamino)-purine. During a 28-day period, the fresh weight of calli increased approximately five times. The same medium without agar was used for establishing cell suspension cultures. Fresh weight of cells increased four times and dry weight approximately doubled during 10 days of culture.Abbreviations 2,4-d 2,4-dichlorophenoxyacetic acid - 2iP 6-(dimethylallylamino)-purine - IBA indolebutyric acid - MS Murashige & Skoog basal salt medium     

The Histology of Seed Development following Crosses between Diploid and Autotetraploid Raspberries (Rubus idaeus L.)

Seed development was studied in crosses made between the diploidforms and autotetraploid mutants of two raspberry (Rubus idaeusL.) cultivars. Drupelet abortion did not appear to be causedby failure of fertilization; it occurred mainly between thefourth and eighth day after pollination, and manifested itselfas simultaneous cessation of growth in all tissues. In healthydrupelets at eight days after pollination, growth appeared tobe proceeding at similar rates in all the tissues except thatthe embryo seemed to be growing faster than the endosperm inthe most advanced crosses. Both diploid and tetraploid ovulesappeared to be more stimulated by pollen from tetraploid plants.Embryo differentiation differed according to the direction ofcross, especially in stunted embryos. Radicles tended to beabnormally large when diploid pollen was used and smaller thanusual with haploid pollen.     

Fruit fate,frugivory, and fruit characteristics: a study of the hawthorn,Crataegus monogyna (Rosaceae)

Summary The fate of fruits from a population of European hawthorn (Crataegus monogyna) in western Oregon, USA, was examined over a two-year period. Only one frugivore, the American robin (Turdus migratorius) foraged on the C. monogyna fruits, making this an unusually straightforward fruit-frugivore system. Dispersal efficiency was low, with an average 21% of seeds being dispersed (carried away from parent plants) each year; the most common fate of fruits was to simply fall. Robins dropped 20% of the fruits that they picked, and defecated/regurgitated 40% of the fruits (seeds) that they swallowed, beneath parent plants. One trait, gruit abundance, strongly affected the probability of bush visitation by robins, bushes with larger fruit displays being preferred. Both absolute dispersal success (number of seeds) and dispersal efficiency (proportion of seeds; success per propagule) were also found to be correlated most strongly with initial fruit abundance. Individual plant fecundity and fruit quality were found to vary little between years; as a result, dispersal efficiencies for individual plants were also annully consistent. Larger (older) plants produced more fruits and therefore had higher fitness. These results suggest that the optimal fruiting strategy for C. monogyna is therefore to get as big as possible as quickly as possible by delaying fruiting until later in life.     

The role of ethylene and cell wall modifying enzymes in raspberry (Rubus idaeus) fruit ripening

This study focuses on four raspberry ( Rubus idaeus ) genotypes from two different genetic backgrounds: cvs Glen Prosen and Glen Clova, bred at the Scottish Crop Research Institute (SCRI) and genotypes bred at Horticulture Research International (HRI), East Malling (EM), EM 4997 and EM 5007. The ripe fruit of each genotype pair were characterised subjectively by raspberry breeders as relatively firm or soft, respectively. Different stages of fruit development from each genotype were used to quantify fruit firmness, rates of ethylene evolution and ripening rate. Penetrometry data confirmed suspected firmness differences. Firmness correlated with rates of ethylene evolution. Rates of ethylene production also correlated with receptacle size. Storage of green fruits in 20 μl l⁻¹ ethylene reduced fruit firmness, enhanced respiration rate and colour (anthocyanin) development and stimulated the development of cell wall hydrolase activities. However, during natural ripening in the field, fruit respiration rate declined, which indicates a non-climacteric ripening pattern. In drupelets, the activities of polygalacturonase (PG), pectin methylesterase (PME), C x -cellulase (C x ) and β -galactosidase ( β -gal.) increased substantially as ripening progressed. More detailed studies with ripe fruit of cv. Glen Clova indicated major isoforms of PG at pIs 3.3, 8.6 and 10.1; of PME at pIs 7.2, 8.5, 8.7, 8.8; of C x at pI 2.4; and of β -gal. at pIs 6.3 and 6.7.     

The Structure of the Fruit of the Red Raspberry (Rubus idaeus L.) in Relation to Abscission

Two modes of cell separation occur within raspberry abscissionzones. In the cortex, fracture of the middle lamella, partialdissolution of the cell wall, and growth stresses in the abscissionzone cause existing intercellular spaces to develop into largecavities. Cells remain intact during this process. In the vascularbundle, dissolution of the middle lamella and the resultantformation of cavities in the cell wall is followed by cell wallbreakdown and cell disintegration. Intracellular membrane systemsappear to be associated with areas of middle lamellar dissolution. Rubus idaeus L., raspberry, fruit structure, abscission     

Chemical composition and ultrastructure of the epicuticular wax in three lines of Brassica napus (L)

The epicuticular wax in three lines of Brassica napus (rape) has been investigated and the detailed chemistry and ultrastructure of the waxes examined. A distinct chemical make-up has been found for all three waxes which is correlated with three distinct crystallite structures. A tentative scheme for classification of Brassica wax mutants is described in which the two newly analysed rape mutants can be placed. Mass spectral analysis of all wax components confirms and extends previous ideas about the chemistry of Brassica waxes.     

Chemical composition and ultrastructure of the epicuticular wax in four mutants of Pisum sativum (L)

The action of mutations affecting the epicuticular wax of Pisum sativum has been investigated at the chemical and ultrastructural level. Upper and lower surfaces of the leaves were found to differ markedly in both ultrastructure and chemistry. Mutations affected primarily either the lower (wa, wb and wsp) or the upper surface (wlo), but some effects of all 4 genes could be seen on both surfaces. Specific biochemical lesions could be implied for wsp and wa but the chemical effects of wb and wlo were more diffuse. Generally a close relation between chemical composition and crystallite form of the wax was evident throughout the work.