Introgression and confirmation of everbearing trait in strawberry (Fragaria x ananassa Duch.)

The present investigation was targeted towards a highly desirable everbearing trait in strawberry (Fragaria × ananassa Duch.) via marker assisted selection while seeing its worldwide commercial applicability through the extended harvest season. The crosses were made between everbearing and june-bearing cultivars to raise the F₁ individuals. Morphological characters (plant, floral, and fruit) were assessed that showed significant differences among the strawberry cultivars. Molecular characterization was carried out between everbearing and non-everbearing cultivars using RAPD and SSR markers. For phenotyping, a chi-square test was performed and revealed that out of all four cross combinations, the best fitted cross found to be in Mendelian segregation ratio (1:1) was ‘Confectura’ × ‘Torrey’ with χ²-value 1.58. Further, the identified polymorphic markers were assessed across the F₁ individuals of cross ‘Confectura’ × ‘Torrey’ for its genotyping. It could be revealed that the targeted everbearing trait is governed by a dominant gene(s) in the subjected strawberry genotypes. Further, the identified polymorphic markers would be successfully employed in DNA fingerprinting of strawberry under various crop improvement programme.Supplementary informationThe online version of this article (10.1007/s12298-020-00916-w) contains supplementary material, which is available to authorized users.     

Metabolic profiling of strawberry (Fragaria x ananassa Duch.) during fruit development and maturation

Strawberry (Fragaria × ananassa Duch), a fruit of economic and nutritional importance, is also a model species for fleshy fruits and genomics in Rosaceae. Strawberry fruit quality at different harvest stages is a function of the fruit's metabolite content, which results from physiological changes during fruit growth and ripening. In order to investigate strawberry fruit development, untargeted (GC-MS) and targeted (HPLC) metabolic profiling analyses were conducted. Principal component analysis (PCA) and partial least squares discriminant analysis (PLS-DA) were employed to explore the non-polar and polar metabolite profiles from fruit samples at seven developmental stages. Different cluster patterns and a broad range of metabolites that exerted influence on cluster formation of metabolite profiles were observed. Significant changes in metabolite levels were found in both fruits turning red and fruits over-ripening in comparison with red-ripening fruits. The levels of free amino acids decreased gradually before the red-ripening stage, but increased significantly in the over-ripening stage. Metabolite correlation and network analysis revealed the interdependencies of individual metabolites and metabolic pathways. Activities of several metabolic pathways, including ester biosynthesis, the tricarboxylic acid cycle, the shikimate pathway, and amino acid metabolism, shifted during fruit growth and ripening. These results not only confirmed published metabolic data but also revealed new insights into strawberry fruit composition and metabolite changes, thus demonstrating the value of metabolomics as a functional genomics tool in characterizing the mechanism of fruit quality formation, a key developmental stage in most economically important fruit crops.     

Protection of strawberry plants (Fragaria ananassa Duch.) against anthracnose disease induced by Azospirillum brasilense

Plant and Soil - Agronomic biofortification of Zn requires an effective Zn application method and efficient Zn utilization by the crops. Various Zn application methods were compared for Zn...     

Polyamines,floral induction and floral development of strawberry (Fragaria ananassa Duch.)

In the short-day plant, strawberry (Fragaria ananassa Duch.), polyamines (putrescine, spermidine and spermine), conjugated spermidine (water-insoluble compounds) and bound amines (putrescine, spermidine, phenylethylamine, 3-hydroxy, 4-methoxyphenylethylamine) accumulated in the shoot tips during floral induction and before floral emergence. Different associations of free amines and conjugated amines were observed during floral induction, as compared with the reproductive phase. During the whole period of floral development, phenylethylamine (an aromatic amine) was the predominant amine, representing 80 to 90% of the total free amine pool. Phenylethylamine conjugates (water-insoluble compounds) were the predominant amides observed prior to fertilization. These substances decreased drastically after fertilization. In vegetative shoot tips from plants grown continously under long days, free polyamines (putrescine, spermidine) and bound polyamines (putrescine, spermidine) were low and no change was observed. Free amines (spermine and phenylethylamine), bound aromatic amines (phenylethylamine, 3-hydroxy, 4-methoxyphenylethylamine), conjugated spermidine and phenylethylamine did not appear. Male-sterile flowers were distinguished by their lack of conjugated spermidine and phenylethyalamine and by a decrease in free phenylethylamine. In normal and sterile strawberry plants -DL-difluoromethylornithine (DFMO), a specific irreversible inhibitor of ornithine decarboxylase (ODC), caused inhibition of flowering and free and polyamine conjugates. When putrescine was added, polyamine titers and flowering were restored. A similar treatment with -DL-difluoromethylarginine (DFMA), a specific, irreversible inhibitor of arginine decarboxylase (ADC), did not affect flowering and polyamine titers. These results suggest that ornithine decarboxylase (ODC) and polyamines are involved in regulating floral initiation in strawberry. The relationship between polyamines, aromatic amines, conjugates, floral initiation and male sterility is discussed.Abbreviations ADC arginine decarboxylase - ODC ornithine decarboxylase - DFMA -DL-difluoromethylarginine - DFMO -DL-difluoromethylornithine - Put putrescine - Spd spermidine - Spm spermine - Phen phenylethylamine - 3H4M Phen 3-hydroxy, 4-methoxyphenylethylamine     

Haploid plant regeneration from anther cultures of three north american cultivars of strawberry (Fragaria x ananassa Duch.)

Summary A study was conducted to maximize plant regeneration frequencies from cultured anthers of Chandler, Honeoye, and Redchief strawberries (Fragaria x ananassa Duch.). A comparison of auxins (IAA, NAA), cytokinins (BA, BPA, KIN) and carbohydrates (sucrose, glucose, maltose) in MS medium showed that the highest shoot regeneration across cultivars (8%) occurred when using a medium containing 2 mg/l IAA, 1 mg/l BA, and 0.2 M glucose. A comparison of MS, NN, and H1 inorganic medium (a new formulation based on the anther culture literature) solidified with either agar or gellan gum and containing IAA, BA, and glucose, showed the highest shoot regeneration across cultivars (19%) when using H1 and gellan gum. Lastly, media containing Fe-EDTA yielded more shoots than media containing Fe-Metalosate, and anthers cultured on Fe-EDTA media in darkness for 30d followed by 30d in white light produced more shoots (16% average regeneration) than those cultured on Fe-EDTA media under white or yellow light (16h photoperiod) for the initial 30d (0.3% and 5% respectively). Plants were acclimated ex vitro where they flowered and set fruit. Chromosome counts of root tip cells confirmed that haploid plants were obtained from all three cultivars.Abbreviations IAA indoleacetic acid - NAA naphthaleneacetic acid - BA 6-benzylaminopurine - BPA N-benzyl-9-(2-tetrahydropyranyl)-adenine - KIN 6-furfurylaminopurine - MS Murashige & Skoog (1962) - NN Nitsch & Nitsch (1969)     

Vermicompost substitution influences growth, physiological disorders, fruit yield and quality of strawberry (Fragaria x ananassa Duch.)

Studies were conducted to determine the effect of vermicompost on growth, physiological disorders, fruit yield and quality of 'Chandler' strawberry. For this, 4 levels of vermicompost (2.5, 5.0, 7.5 and 10.0tha(-1)) were supplemented with inorganic fertilizers to balance fertilizer requirement of strawberry under semi-arid region of northern India. The vermicompost was incorporated into top 10cm layer of soil, which was supplemented on the basis of chemical analysis, with amount of inorganic N, P, K fertilizer calculated to equalize the recommended dose of nutrients. Vermicompost application increased plant spread (10.7%), leaf area (23.1%) and dry matter (20.7%), and increased total fruit yield (32.7%). Substitution of vermicompost drastically reduced the incidence of physiological disorders like albinism (16.1-4.5%); fruit malformation (11.5-4.0%) and occurrence of grey mould (10.4-2.1%) in strawberry indicating that vermicompost had significant role in reducing nutrient-related disorders and disease like Botrytis rot, and thereby increasing the marketable fruit yield up to 58.6% with better quality parameters. Fruit harvested from plant receiving vermicompost were firmer, have higher TSS, ascorbic acid content and lower acidity, and have attractive colour. All these parameters appeared to be dose dependent and best results were achieved @ 7.5tha(-1), however, beyond this dose of vermicompost, there was not significant influence on these parameters.     

In vitro selection in resistance breeding of strawberry (Fragaria x ananassa duch.)

Genetic resistance to pathogenetic soil-borne fungus Verticillium dahliae Kleb. was examined in two strawberry somaclones. Strawberry somaclones were obtained in sterile culture from runner tips of cultivars 'Merton Dawn' and 'Selva'. In vitro selection was performed with the use of homogenate of liquid cultures of Verticillium dahliae. Microplants of both somaclones were inoculated at stage of 4. Leaves. Disease symptoms were observed at 15., 30., 45., 60. and 75. days post inoculation. Extent of leaf chlorosis was rated on a scale of 0-4. Under the controlled in vitro culture conditions a different response to infection by this pathogenic fungus was observed. After 75. days post inoculation the contribution of necrotic plants in somaclone of 'Merton Dawn' reached the value of 76%, whereas in somaclone of 'Selva' this value reached 86%. In control somaclones of 'Merton Dawn' and 'Selva' the contribution of necrotic plants after 75. days post mock-inoculation with sterile distilled water reached the considerably lower value of 13%. These results revealed that somaclone of 'Merton Dawn' was more genetically resistant to infection by V. dahliae than somaclone of 'Selva'. The observed response to in vitro infection caused by Verticillium dahliae in examined somaclones was similar in comparison with original cultivars. Furthermore, somaclonal variation induced in tissue cultured strawberry was sufficient to select variants that showed enhanced genetic resistance to Verticillium wilt caused by V. dahliae. In vitro selection can be efficiently used as an alternative program to conventional resistance breeding in strawberry.     

Response of six strawberry (Fragaria x ananassa Duch.) cultivars to the root lesion nematode (Pratylenchus penetrans Filipjev and Schurmanns Stekhoven)

In the Philippines, strawberry is grown only in Benguet Province because of its unique climatic conditions. It has been a lucrative source of income for Benguet farmers and adds to the revenue of Benguet Province. The root lesion nematode, Pratylenchus penetrans is an economically important pest of strawberry in the area. It can cause substantial losses to strawberry growers, both by reducing vegetative plant growth and by reducing strawberry yields. The nematode has a very wide host range and hence, is not readily controlled by crop rotation. An alternative approach which growers may wish to consider trying is planting of strawberry varieties which are either resistant or tolerant to this nematode. The relative susceptibility/tolerance of six strawberry cultivars to the root lesion nematode, P. penetrans was evaluated under greenhouse and field conditions. Inoculation of 500 nematodes/pot did not significantly affect the fresh top weight, fresh root weight, and yield of strawberry cultivars Festival, Whitney, Winterdawn, Earlibrite, and Camarosa. The said cultivars had significantly higher number of nematodes recovered from the roots. On the other hand, the highest strawberry yield was recorded in cv Sweet Charlie, however, this was significantly reduced by nematode inoculation .Surprisingly, the number of nematodes recovered from the roots of this cultivar was significantly the lowest among the cultivars tested. Results of the field experiment showed that strawberry cv Sweet Charlie gave the highest marketable yield which was significantly different from the rest of the cultivars tested. This was followed by Festival, and Earlibrite. On the other hand, Camarosa and Whitney gave significantly lower yield than the above cultivars but significantly higher than Winterdawn. In terms of nematodes recovered from the roots, the highest was noted in Whitney, followed by Sweet Charlie and Earlibrite. The lowest was obtained from Camarosa, followed by Festival and Winterdawn. Based on the results of the greenhouse experiment, Festival, Whitney, Winterdawn, Earlibrite and Camarosa can be considered tolerant while Sweet Charlie was slightly susceptible to P. penetrans. However, based on the field trial, Sweet Charlie, Festival and Earlibrite were tolerant while Whitney, Camarosa and Winterdawn were slightly susceptible.     

Growth of hairy root cultures of strawberry (Fragaria ¥ ananassa Duch.) in three different types of bioreactors

Hairy roots of strawberry were cultivated in three different types of bioreactors: an air-sparged bioreactor (control), a droplet bioreactor and a mist bioreactor. The highest biomass yields (3.7 g dry wt/l) were achieved in the air-sparged and in the mist bioreactor. In the droplet bioreactor the cultivation medium was insufficiently atomized into droplets and nutrient uptake and growth were slower due to uneven wetting of hairy roots.     

Fruit Set and Growth in Strawberry, Fragaria x ananassa Duch.

The application of GA₃in aqueous solution to leaves or flowersof hermaphrodite cultivars of strawberry, Redgauntlet and Rabunda,prevented growth of the receptacle despite hand pollination.This inhibitory effect occurred only when GA₃ was applied priorto anthesis. Although viable pollen was produced and germinatedto grow down the styles of treated plants, no seeds were formed.Receptacle growth failed underneath the unfertilized carpels,but the basal region devoid of carpels enlarged and ripened.The effect of GA₃ was the same in vivo and for flowers grownin vitro. ABA and BAP also inhibited growth of pollinated flowersin vitro, but neither substance stimulated growth of the baseof the receptacle. 2-NOA stimulated receptacle growth of pollinatedflowers but did not overcome the inhibitory effect of GA₃. Removal of fertile carpels 9 days after pollination preventedfurther receptacle growth. GA₃ treatment of the bare receptaclere-started growth but was less effective than 2-NOA. No growth substance treatment induced parthenocarpic developmentin these cultivars when unopened buds were emasculated and culturedwithout pollination, although GA₃ induced some swelling of thereceptacle base. Fragaria x ananassa Duch., strawberry fruit set, fruit growth, growth regulators     

Infection of hairy roots of strawberry (Fragaria x Ananassa Duch.) with arbuscular mycorrhizal fungus

Summary Hairy root cultures of strawberry (Fragaria x ananassa Duch.) were induced with the Agrobacterium rhizogenes strain A4. Cultures were maintained on B50 medium but could also grow on a minimal medium, which did not inhibit the growth of arbuscular mycorrhizal fungi. The growth and nutrient uptake were characterized in shake flasks and in a bioreactor. Spores of the native Finnish arbuscular mycorrhizal fungus Glomus fistulosum V128 were used to infect strawberry (Fragaria x ananassa Duch. Senga Sengana) hairy roots in vitro. During cultivation, vegetative spore formation was observed. At the end of the cultivation, hyphae and arbuscules were observed in the stained roots.Abbreviations AM arbuscular mycorrhiza - AMF arbuscular mycorrhizal fungus     

Multi-substrate flavonol O-glucosyltransferases from strawberry (Fragaria x ananassa) achene and receptacle

In an effort to characterize fruit ripening-related genes functionally, two glucosyltransferases, FaGT6 and FaGT7, were cloned from a strawberry (Fragaria x ananassa) cDNA library and the full-length open reading frames were amplified by rapid amplification of cDNA ends. FaGT6 and FaGT7 were expressed heterologously as fusion proteins in Escherichia coli and target protein was purified using affinity chromatography. Both recombinant enzymes exhibited a broad substrate tolerance in vitro, accepting numerous flavonoids, hydroxycoumarins, and naphthols. FaGT6 formed 3-O-glucosides and minor amounts of 7-O-, 4'-O-, and 3'-O-monoglucosides and one diglucoside from flavonols such as quercetin. FaGT7 converted quercetin to the 3-O-glucoside and 4'-O-glucoside and minor levels of the 7- and 3'-isomers but formed no diglucoside. Gene expression studies showed that both genes are strongly expressed in achenes of small-sized green fruits, while the expression levels were generally lower in the receptacle. Significant levels of quercetin 3-O-, 7-O-, and 4'-O-glucosides, kaempferol 3-O- and 7-O-glucosides, as well as isorhamnetin 7-O-glucoside, were identified in achenes and the receptacle. In the receptacle, the expression of both genes is negatively controlled by auxin which correlates with the ripening-related gene expression in this tissue. Salicylic acid, a known signal molecule in plant defence, induces the expression of both genes. Thus, it appears that FaGT6 and FaGT7 are involved in the glucosylation of flavonols and may also participate in xenobiotic metabolism. The latter function is supported by the proven ability of strawberries to glucosylate selected unnatural substrates injected in ripe fruits. This report presents the first biochemical characterization of enzymes mainly expressed in strawberry achenes and provides the foundation of flavonoid metabolism in the seeds.     

A fruit-specific and developmentally regulated endopolygalacturonase gene from strawberry (Fragaria x ananassa cv. Chandler)

A fruit-specific and developmentally regulated polygalacturonase gene (spG gene) from strawberry (Fragaria x ananassa cv. Chandler) has been cloned and characterized at a molecular and physiological level. Comparison analysis of the corresponding deduced sPG protein have shown that this strawberry gene is similar to Clade A endopolygalacturonase genes. Moreover, the spatio-temporal and hormonal gene expression pattern suggests a close relationship between the expression of this gene and the onset of the strawberry fruit ripening process and agrees with that of the production of oligosaccharins which have already been described as active molecules involved in fruit ripening. The results are discussed in terms of a putative role of this enzyme in the release of oligosaccharins from the strawberry fruit cell wall.     

RNAi-induced silencing of gene expression in strawberry fruit (Fragaria x ananassa) by agroinfiltration: a rapid assay for gene function analysis

Intron-containing constructs encoding self-complementary 'hairpin' RNA (ihpRNA) have the potential to efficiently silence genes in a range of plant species. In this study we demonstrate the silencing of a ripening-related chalcone synthase (CHS) gene in strawberry fruits (Fragaria x ananassa cv. Elsanta) by a construct (ihpRNA) containing the partial sense and corresponding antisense sequences of CHS separated by an intron obtained from a F. x ananassa quinone oxidoreductase gene. An Agrobacterium strain carrying a T-DNA expressing the ihpRNA transgene was injected with a syringe into the receptacles of growing fruits still attached to the plant about 14 days after pollination. As a consequence of the reduced levels of CHS mRNA and enzymatic CHS activity, the levels of anthocyanins were downregulated and precursors of the flavonoid pathway were shunted to the phenylpropanoid pathway leading to a large increases in levels of (hydroxy) cinnamoyl glucose esters. We anticipate that this technique in combination with metabolite profiling analysis will be useful for studying the function of unknown genes during the development and ripening of strawberry fruit.     

Cloning,expression and immunolocalization pattern of a cinnamyl alcohol dehydrogenase gene from strawberry (Fragaria x ananassa cv. Chandler)

Cinnamyl alcohol dehydrogenase (CAD; EC 1.1.1.195) catalyses the conversion of p-hydroxy-cinnamaldehydes to the corresponding alcohols and is considered a key enzyme in lignin biosynthesis. By a differential screening of a strawberry (Fragariax ananassa cv. Chandler) fruit specific subtractive cDNA library, a full-length clone corresponding to a cad gene was isolated (Fxacad1). Northern blot and quantitative real time PCR studies indicated that the strawberry Fxacad1 gene is expressed in fruits, runners, leaves, and flowers but not in roots. In addition, the gene presented a differential expression in fruits along the ripening process. Moreover, by screening of a strawberry genomic library a cad gene was isolated (Fxacad2). Similar to that found in other cad genes from higher plants, this strawberry cad gene is structured in five exons and four introns. Southern blot analyses suggest that, probably, a small cad gene family exists in strawberry. RT-PCR studies indicated that only the Fxacad1 gene was expressed in all the fruit ripening stages and vegetative tissues analysed. The Fxacad1 cDNA was expressed in E. coli cells and the corresponding protein was used to raise antibodies against the strawberry CAD polypeptide. The antibodies obtained were used for immunolocalization studies. The results showed that the CAD polypeptide was localized in lignifying cells of all the tissues examined (achenes, fruit receptacles, runners, leaves, pedicels, and flowers). Additionally, the cDNA was also expressed in yeast (Pichia pastoris) as an extracellular protein. The recombinant protein showed activity with the characteristic substrates of CAD enzymes from angiosperms, indicating that the gene cloned corresponds to a CAD protein.     

草莓果实MADS-box基因保守片段的克隆与序列分析

根据多种植物MADS-box基因保守区序列设计简并引物,应用PCR技术从草莓绿果中分高出33条MADS-box基因cDNA片段.序列分析表明,这些片段长度在137 - 146 bp之间,包含基因起始密码子.推导的氨基酸序列与已登录的草莓的MADS-box基因同源性超过87％.推导的氨基酸序列与已知的革莓和其他物种调控果实发育成熟的MADs-box基因以及拟南芥的MADS-box家族基因进行系统发育分析,可将这些基因片段分科归入拟南芥MADS-box基因不同亚家族中,证明草莓果实中存在各类MADS-box家族基因,克隆的部分片段可能参与调控果实发育和成熟软化的调节.