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Background The plant hormone auxin is widely involved in plant growth, development, and morphogenesis, and the TIR1/AFB and AUX/IAA proteins are closely linked to rapid auxin response and signal transmission. However, their evolutionary history, historical patterns of expansion and contraction, and changes in interaction relationships are still unknown. Results Here, we analyzed the gene duplications, interactions, and expression patterns of TIR1/AFBs and AUX/IAAs to understand their underlying mechanisms of evolution. The ratios of TIR1/AFBs to AUX/IAAs range from 4:2 in Physcomitrium patens to 6:29 in Arabidopsis thaliana and 3:16 in Fragaria vesca. Whole-genome duplication (WGD) and tandem duplication have contributed to the expansion of the AUX/IAA gene family, but numerous TIR1/AFB gene duplicates were lost after WGD. We further analyzed the expression profiles of TIR1/AFBs and AUX/IAAs in different tissue parts of Physcomitrium patens, Selaginella moellendorffii, Arabidopsis thaliana and Fragaria vesca, and found that TIR1/AFBs and AUX/IAAs were highly expressed in all tissues in P. patens, S. moellendorffii. In A. thaliana and F. vesca, TIR1/AFBs maintained the same expression pattern as the ancient plants with high expression in all tissue parts, while AUX/IAAs appeared tissue-specific expression. In F. vesca, 11 AUX/IAAs interacted with TIR1/AFBs with different interaction strengths, and the functional specificity of AUX/IAAs was related to their ability to bind TIR1/AFBs, thus promoting the development of specific higher plant organs. Verification of the interactions among TIR1/AFBs and AUX/IAAs in Marchantia polymorpha and F. vesca also showed that the regulation of AUX/IAA members by TIR1/AFBs became more refined over the course of plant evolution. Conclusions Our results indicate that specific interactions and specific gene expression patterns both contributed to the functional diversification of TIR1/AFBs and AUX/IAAs.

BACKGROUND: SQUAMOSA promoter binding protein-like (SPL) proteins are a class of plant-specific transcription factors that play important roles in plant development. However, the majority of SPL genes in strawberry are functionally uncharacterized. OBJECTIVE: To understand the biological functions and elucidate the molecular regulatory mechanisms of SPL genes in strawberry development. METHODS: The FvSPL1 gene from woodland strawberry was cloned and the phylogenetic tree was analyzed using bioinformatics methods. Subcellular localization, transcriptional activity, DNA binding ability and regulatory mechanisms of FvSPL1 were analyzed through biochemical and genetic approaches. RESULTS: Phylogenetic analysis with SPL genes from Arabidopsis, tomato and chrysanthemum indicated that FvSPL1 clustered in the same group as those of the miR156 target site located at the 3′-untranslated region. Further biochemical analysis indicated that FvSPL1 was exclusively localized in the nucleus. Electrophoretic mobility shift assay demonstrated that FvSPL1 could specifically recognize the GTAC motif. Transcriptional activity analysis showed that FvSPL1 is a transcriptional activator that could activate the expression of the FvAP1 gene. Finally, transgenic Arabidopsis overexpressing the FvSPL1 gene exhibited significantly early flowering. Taken together, our study indicated that FvSPL1, similar to its orthologs in Arabidopsis, mainly functions in regulating plant flowering. CONCLUSIONS: These findings provide insight into the mechanism of flowering in strawberry, and contribute to the understanding of strawberry flowering time manipulation which will facilitate the molecular breeding of strawberries.

Presence and absence polymorphisms (PAPs) exist extensively and have been investigated in different organisms. However, PAPs have rarely been detected between strawberry species at the genome level. This study identified the presence and absence genes (P/A genes) between wild strawberry species (Fragaria vesca) and octoploid cultivated species (F. × ananassa) under a relatively strict criterion. In total, 333 P/A genes present in the wild strawberry but absent in the cultivated strawberry were detected. Of the P/A genes, 91.89% (306/333) were single genes, and only 8.11% were confirmed as multi-genes. The majority of the identified P/A genes in Fragaria were generated by tandem duplications. The P/A genes were unevenly distributed on the seven chromosomes of woodland strawberry, and they clustered preferentially near the telomeric regions of the chromosomes. The P/A genes tended to encode proteins with domains closely associated with responses to varying ecological factors, such as PPR, Protein kinases (PKs), NB-ARC, F-box and EF-hand domains. This indicated that the P/A genes were associated with coping with biotic and abiotic stresses to improve the adaptability of plants to changing environments.

Fragaria vesca, commonly known as wild or woodland strawberry, is the most widely distributed diploid Fragaria species and is native to Europe and Asia. Because of its small plant size, low heterozygosity, and relative ease of genetic transformation, F. vesca has been a model plant for fruit research since the publication of its Illumina-based genome in 2011. However, its genomic contribution to octoploid cultivated strawberry remains a long-standing question. Here, we de novo assembled and annotated a telomere-to-telomere, gap-free genome of F. vesca ‘Hawaii 4’, with all seven chromosomes assembled into single contigs, providing the highest completeness and assembly quality to date. The gap-free genome is 220 785 082 bp in length and encodes 36 173 protein-coding gene models, including 1153 newly annotated genes. All 14 telomeres and seven centromeres were annotated within the seven chromosomes. Among the three previously recognized wild diploid strawberry ancestors, F. vesca, F. iinumae, and F. viridis, phylogenomic analysis showed that F. vesca and F. viridis are the ancestors of the cultivated octoploid strawberry F. × ananassa, and F. vesca is its closest relative. Three subgenomes of F. × ananassa belong to the F. vesca group, and one is sister to F. viridis. We anticipate that this high-quality, telomere-to-telomere, gap-free F. vesca genome, combined with our phylogenomic inference of the origin of cultivated strawberry, will provide insight into the genomic evolution of Fragaria and facilitate strawberry genetics and molecular breeding.

Genes encoding VQ motif-containing (VQ) transcriptional regulators and WRKY transcription factors can participate separately or jointly in plant growth, development, and abiotic and biotic stress responses. In this study, 222 VQ and 645 WRKY genes were identified in six Prunus species. Based on phylogenetic tree topologies, the VQ and WRKY genes were classified into 13 and 32 clades, respectively. Therefore, at least 13 VQ gene copies and 32 WRKY gene copies were present in the genome of the common ancestor of the six Prunus species. Similar small Ks value peaks for the VQ and WRKY genes suggest that the two gene families underwent recent duplications in the six studied species. The majority of the Ka/Ks ratios were less than 1, implying that most of the VQ and WRKY genes had undergone purifying selection. Pi values were significantly higher in the VQ genes than in the WRKY genes, and the VQ genes therefore exhibited greater nucleotide diversity in the six species. Forty-one of the Prunus VQ genes were predicted to interact with 44 of the WRKY genes, and the expression levels of some predicted VQ-WRKY interacting pairs were significantly correlated. Differential expression patterns of the VQ and WRKY genes suggested that some might be involved in regulating aphid resistance in P. persica and fruit development in P. avium.

The various monochromatic Light Emitting Diode (LED) lights are widely used in growth facility for cultivating various plants, particularly horticultural crops because of their higher luminous efficiency, lower radiation and power consumption than the traditional white fluorescent lamp light. However, considerable inconsistent effects have been reported in literature. We conducted a meta-analysis to assess the effects of different colors of LED light on plant growth, development and various traits. Compared to the light from white fluorescent lamps, the red LED light significantly changed 4 out 26 plant characteristics by at least 37%, and blue LED light significantly increased 5 of 26 assessed characteristics by 37% or more. The combination of red/blue LED lights only significantly increased dry weight by 161% among 25 plant characteristics analyzed. Compared to the white LED light, red LED light significantly decreased 2 of 9 plant characteristics by at least 36%, and blue LED light significantly decreased only 1 of 9 plant characteristics, total chlorophyll content, by 42%. In the moderators analyzed, plant taxonomic families significantly influenced the effects of LED lights on shoot dry weight, and plant life cycles and plant taxonomic families significantly affected the effect on stomatal conductance. Through systematic meta-analysis, we found that the effect of LED on plant growth and quality traits was species-specific, and the effect was affected by the cultivation conditions. Therefore, we suggest that researchers be more targeted to experiment, and collect traits associated with practical production, especially related to the quality of product data, such as carotenoids, anthocyanin and other antioxidant compounds. This article is to provide more data with practical application, guide the application of LED in horticultural plant factory.

Background GRAS genes formed one of the important transcription factor gene families in plants, had been identified in several plant species. The family genes were involved in plant growth, development, and stress resistance. However, the comparative analysis of GRAS genes in Rosaceae species was insufficient. Results In this study, a total of 333 GRAS genes were identified in six Rosaceae species, including 51 in strawberry (Fragaria vesca), 78 in apple (Malus domestica), 41 in black raspberry (Rubus occidentalis), 59 in European pear (Pyrus communis), 56 in Chinese rose (Rosa chinensis), and 48 in peach (Prunus persica). Motif analysis showed the VHIID domain, SAW motif, LR I region, and PFYRE motif were considerably conserved in the six Rosaceae species. All GRAS genes were divided into 10 subgroups according to phylogenetic analysis. A total of 15 species-specific duplicated clades and 3 lineage-specific duplicated clades were identified in six Rosaceae species. Chromosomal localization presented the uneven distribution of GRAS genes in six Rosaceae species. Duplication events contributed to the expression of the GRAS genes, and Ka/Ks analysis suggested the purification selection as a major force during the evolution process in six Rosaceae species. Cis-acting elements and GO analysis revealed that most of the GRAS genes were associated with various environmental stress in six Rosaceae species. Coexpression network analysis showed the mutual regulatory relationship between GRAS and bZIP genes, suggesting the ability of the GRAS gene to regulate abiotic stress in woodland strawberry. The expression pattern elucidated the transcriptional levels of FvGRAS genes in various tissues and the drought and salt stress in woodland strawberry, which were verified by RT-qPCR analysis. Conclusions The evolution and functional analysis of GRAS genes provided insights into the further understanding of GRAS genes on the abiotic stress of Rosaceae species.

BACKGROUND: Grapevine was one of the most important perennial fruit crops worldwide. Acyl-CoA-binding proteins (ACBPs) in eudicots and monocots show conservation in an acyl-CoA-binding domain (ACB domain) which binds acyl-CoA esters. OBJECTIVE: The information and data provided in the present study contributes to understand the evolutionary processes and potential functions of this gene family in grapevine growth and development, and responses to abiotic stress. METHODS: Using the complete grapevine genome sequences, we investigated the number grapevine ACBP genes, the exon-intron structure, phylogenetic relationships and synteny with the Arabidopsis ACBP gene family. Furthermore, the expression profiles of VvACBP genes based on public microarray data in different tissues, and the expression patterns responding to different exogenous hormones as well as abiotic and biotic stresses were presented. The qRT-PCR was used to verify the microarray data under drought stress treatments. Finally, the leaf relative water content (RWC), leaf chlorophyll content, and enzymatic activities were measured to further examine the tolerance to drought stress in grapevine. RESULTS: The six grapevine ACBPs were identified. Their distribution into various groups differed from Arabidopsis and rice. Synteny analysis demonstrated that several VvACBP genes were found in corresponding syntenic blocks of Arabidopsis, suggesting that these genes arose before the divergence of the respective lineages. Sequence alignment and structural annotation provided an overview of variations that might contribute to functional divergence from Arabidopsis ACBPs. Expressional analyses suggested that both conserved and variant biological functions exist in ACBPs across different species. The expression pattern of these genes were similar in the microarray and qRT-PCR analyses. Gene structure organization and expression characteristics of VvACBPs resembled those of their Arabidopsis orthologous, although species-specific differences also exist. Differential regulation of genes suggested functional diversification among isoforms. The biochemical and physiological data showed the tolerance to drought stress of grapevine. CONCLUSIONS: These findings provided insight into evolution of ACBP gene family in plants and a solid foundation for a deeper understanding of the complex molecular responses of grapevine to stress.

There is an increasing demand for fresh fruits such as strawberry enriched in antioxidants and vitamins. Strawberry fruit quality is highly affected by genotype and environmental conditions. In thi...

Apple blotch caused by Alternaria alternata apple pathotype is a severe disease of apple (Malus ×domestica Borkh) occurring throughout the world, especially in eastern Asia. Phenotypic and genetic information about resistance/susceptibility of apple germplasm to this disease will be extremely valuable for selecting and developing new disease resistant cultivars. In this study, 110 apple cultivars obtained from the USDA apple germplasm in Geneva, NY, were evaluated for their resistance/susceptibility to apple blotch by field surveys, and inoculation of detached leaves with a suspension of germinated conidia of A. alternata apple pathotype. Disease incidence were different among the cultivars and categorized into resistant (R), moderately resistant (MR), or susceptible (S). Two molecular markers, S428, a random amplified polymorphic DNA (RAPD) marker associated with disease resistance, and a simple sequence repeat (SSR or microsatellite) marker CH05g07, linked to susceptibility were used to correlate the phenotypes expressed in field surveys and laboratory inoculations. The detection using either the S428 marker or the CH05g07 marker in 50 common breeding cultivars was consistent with R or S traits except for ‘Bisbee’ and ‘Priscilla’. These two cultivars were MR to apple blotch through phenotyping. However, SSR markers were detected, but RAPD markers were not and therefore were considered susceptible. Combined with the record of resistance to fire blight from Germplasm Resources Information Network (GRIN), ‘Dayton’, ‘Mildew Immune Seedling’, ‘Puregold’, and ‘Pumpkin Sweet’ were highly resistant to both diseases and considered as the best choices of parents for stacking resistance to multiple diseases in breeding program.

Overexpression of grapevine VvABF2 gene could enhance osmotic stress tolerance in Arabidopsis thaliana but fully required for ABA signaling. The abscisic acid (ABA)-dependent AREB/ABF-SnRK2 pathway has been demonstrated to play a pivotal role in response to osmotic stress in model plants. However, its function in other specific species, for example grapevine, has not been fully characterized. In this study, grapevine (Vitis vinifera L.) ABSCISIC ACID RESPONSE ELEMENT-BINDING FACTOR2 (VvABF2), a homologous gene of AREB/ABFs form Arabidopsis, was isolated and constitutively expressed in Arabidopsis under the control of the cauliflower mosaic virus 35S promoter. The VvABF2 transgenic Arabidopsis plants showed to be more sensitive to exogenous ABA compared to wild type plants and exhibited significant osmotic tolerance, like polyethylene glycol (PEG) and drought but fully required ABA for signaling. This fact was further confirmed by its downstream gene expression assays. In addition, the determination of ROS antioxidant enzymes (including SOD, POD and CAT) and the MDA of transgenic lines indicated that overexpression of VvABF2 in Arabidopsis significantly increased ROS scavenging ability and thereby reduced the cell membrane damage, which might be ABA-independent. Our results provide evidence that VvABF2 has a similar function to the Arabidopsis homolog in response to osmotic stresses, and that there is a similar ancestral function of this gene in ABA-dependent response to stresses in grapevine.

SQUAMOSA promoter binding protein-like (SPL) proteins is a class of plant specific transcription factors that play important roles during plant development. However, the majority of SPL genes in strawberry are functionally uncharacterized. In this study, three SPL genes, i.e. FvSPL1, FvSPL2, and FvSPL11 (FvSPL1/2/11), from woodland strawberry were cloned and characterized. Phylogenetic analysis with SPL genes from Arabidopsis, tomato and chrysanthemum indicated that FvSPL1/2/11 were clustered into the same group with those of miR156 target site located at the 3’-untranslated region (UTR). Further biochemical analysis indicated that FvSPL1 was exclusively localized in the nucleus. Electrophoretic mobility shift assay demonstrated FvSPL1 could specifically recognized the GTAC motif. Transcriptional activity analysis showed FvSPL1 was a transcriptional activator that could activate the expression of FvAP1 gene. Finally, all of the transgenic Arabidopsis that overexpression the three FvSPL genes were exhibited significantly early flowering phenotype. Taken together, our study indicated that FvSPL1/2/11 similar to their orthologs in Arabidopsis mainly functions in regulating plant flowering. These results enriched our understanding to the functions of SPL genes in strawberry and might be utilized for strawberry flowering time manipulation in the future.

In this study, genome-wide identification, phylogenetic relationships, duplication time and selective pressure of the NBS-LRR genes, an important group of plant disease-resistance genes (R genes), were performed to uncover their genetic evolutionary patterns in the six Prunus species. A total of 1946 NBS-LRR genes were identified; specifically, 589, 361, 284, 281, 318, and 113 were identified in Prunus yedoensis, P. domestica, P. avium, P. dulcis, P. persica and P. yedoensis var. nudiflora, respectively. Two NBS-LRR gene subclasses, TIR-NBS-LRR (TNL) and non-TIR-NBS-LRR (non-TNL), were also discovered. In total, 435 TNL and 1511 non-TNL genes were identified and could be classified into 30/55/75 and 103/158/191 multi-gene families, respectively, according to three different criteria. Higher Ks and Ka/Ks values were detected in TNL gene families than in non-TNL gene families. These results indicated that the TNL genes had more members involved in relatively ancient duplications and were affected by stronger selection pressure than the non-TNL genes. In general, the NBS-LRR genes were shaped by species-specific duplications, and lineage-specific duplications occurred at recent and relatively ancient periods among the six Prunus species. Therefore, different duplicated copies of NBS-LRRs can resist specific pathogens and will provide an R-gene library for resistance breeding in Prunus species.

Background In plant genomes, high proportions of duplicate copies reveals that gene duplications play an important role in the evolutionary processes of plant species. A series of gene families under positive selection after recent duplication events in plant genomes indicated the evolution of duplicates driven by adaptive evolution. However, the genome-wide evolutionary features of young duplicate genes among closely related species are rarely reported. Results In this study, we conducted a systematic survey of young duplicate genes at genome-wide levels among six Rosaceae species, whose whole-genome sequencing data were successively released in recent years. A total of 35,936 gene families were detected among the six species, in which 60.25% were generated by young duplications. The 21,650 young duplicate gene families could be divided into two expansion types based on their duplication patterns, species-specific and lineage-specific expansions. Our results showed the species-specific expansions advantaging over the lineage-specific expansions. In the two types of expansions, high-frequency duplicate domains exhibited functional preference in response to environmental stresses. Conclusions The functional preference of the young duplicate genes in both the expansion types showed that they were inclined to respond to abiotic or biotic stimuli. Moreover, young duplicate genes under positive selection in both species-specific and lineage-specific expansions suggested that they were generated to adapt to the environmental factors in Rosaceae species.

Botrytis cinerea, the causal agent of gray mold disease, is regarded as the most impactful postharvest fungal pathogen, as it causes serious losses in fresh fruits, ornamentals, and vegetables worldwide. To understand the molecular defense mechanisms of the affected plants, we analyzed transcriptome using RNA sequencing (RNA-seq) of mature strawberry fruit that were infected by B. cinerea at 12, 24, and 48 h. Overall, 18 cDNA libraries were produced generating 923,482,642,150 base pair reads. A total of 1459 differentially expressed genes (DEGs) were identified (≥2.0-fold change and FDR ≤ 0.05), among which only 26 were down-regulated. The majority of changes in gene expression occurred 24 h after inoculation, before significant lesion development, suggesting that this is an important stage during B. cinerea infection. Further analysis revealed that the DEGs were predominately involved in pathogen recognition (pattern recognition receptors), signaling transduction (MAPK cascades, CDPKs), defense response (transcription factors, secondary metabolism biosynthesis, pathogenis-related), as well as different sorts of transporter encoding genes. These data provided a global insight into mature strawberry fruit-B. cinerea interaction and might be used for fruit postharvest pathogens and diseases management.

The ABA/water stress/ripening-induced (ASR) gene family exists universally in higher plants, and many ASR genes are up-regulated during periods of environmental stress and fruit ripening. Although a considerable amount of research has been performed investigating ASR gene response to abiotic stresses, relatively little is known about their roles in response to biotic stresses. In this report, we identified five ASR genes in apple (Malus × domestica) and explored their phylogenetic relationship, duplication events, and selective pressure. Five apple ASR genes (Md-ASR) were divided into two clades based on phylogenetic analysis. Species-specific duplication was detected in M. domestica ASR genes. Leaves of ‘Golden delicious’ and ‘Starking’ were infected with Alternaria alternata f. sp. mali, which causes apple blotch disease, and examined for the expression of the ASR genes in lesion areas during the first 72 h after inoculation. Md-ASR genes showed different expression patterns at different sampling times in ‘Golden delicious’ and ‘Starking’. The activities of stress-related enzymes, peroxidase (POD), superoxide dismutase (SOD), catalase (CAT), phenylalanine ammonia lyase (PAL), and polyphenoloxidase (PPO), and the content of malondialdehyde (MDA) were also measured in different stages of disease development in two cultivars. The ASR gene expression patterns and theses physiological indexes for disease resistance suggested that Md-ASR genes are involved in biotic stress responses in apple.

Pyracantha coccinea is a thorny evergreen shrub native to southeast Europe to southeast Asia. It is a popular ornamental plant because of its showy bright red fruits and small white flowers. However, in vitro vegetative propagation of P. coccinea has not been studied. Nodal segments with one or two axillary buds (1 to 1.5 cm in length) were cut and disinfected in a solution of 0.1% (v/v) mercuric chloride (HgCl2) for 5 minutes, and proliferated on Murashige and Skoog (MS) basal medium supplemented with various concentrations 6-benzylaminopurine (6-BA). After 4 weeks, newly formed shoots were transferred to proliferation and rooting media containing various concentrations of indole-3-butyric acid (IBA). Establishment of axillary buds was significantly better with an establishing rate of 67% on basal MS medium augmented with 6.6 µm 6-BA. The best medium for proliferation of shoots was three-fourth basal MS supplemented with 1.5 µm IBA, with a proliferation rate of 3.4 axillary bud. The optimum rooting medium was one-fourth MS basal medium containing 93 µm IBA. Rooting of shoots was as much as 77%. Rooted plantlets were transferred to pots containing vermiculite:perlite:peat (6:1:2) and acclimatized to ambient greenhouse conditions with a 95% survival rate. This protocol can be used for in vitro propagation of P. coccinea.

Propagation of vegetative cuttings is the most efficient and cost-effective method for producing homogeneous horticultural plants and forest trees when apomictic seed is unavailable. This method depends on the formation of adventitious roots (ARs) in stem or leaf cuttings. Improved understanding of the molecular mechanisms that regulate AR development could enhance the application of this technique. In this article, we review the physiological and developmental processes that regulate AR formation. We map gene function in lateral root development to transcriptomic analyses of the stages of AR formation to reveal potential gene function controlling this important process. We then synthesize a model of the regulatory network controlling AR formation that includes the unique aspects of AR formation in woody plants. Finally, we provide a perspective and suggest future directions for elucidation of molecular processes that regulate AR development.

The basic leucine zipper (bZIP) transcription factors are the most diverse members of dimerizing transcription factors. In the present study, 50, 116, and 47bZIPgenes were identified inMalus domestica(apple),Prunus persica(peach), andFragaria vesca(strawberry), respectively. Species-specific duplication was the main contributor to the large number ofbZIPsobserved in apple. After WGD in apple genome, orthologousbZIPgenes corresponding to strawberry on duplicated regions in apple genome were retained. However, in peach ancestor, these syntenic regions were quickly lost or deleted. Maybe the positive selection contributed to the expansion of clade S to adapt to the development and environment stresses. In addition, purifying selection was mainly responsible forbZIPsequence-specific DNA binding. The analysis of orthologous pairs between chromosomes indicates that these orthologs derived from one gene duplication located on one of the nine ancient chromosomes in the Rosaceae. The comparative analysis ofbZIPgenes in three species provides information on the evolutionary fate ofbZIPgenes in apple and peach after they diverged from strawberry.

VQ motif-containing proteins play crucial roles in plant growth, development, and stress responses. However, no information of VQ motif-containing proteins has been studied at the microevolutionary level in species of Fragaria. In this study, a total of 19, 21, 23, 23, 23, and 25 genes containing the VQ motif were identified from the genomes of F. nipponica, F. iinumae, F. orientalis, F. vesca, F. nubicola, and F. x ananassa, respectively. We classified the VQ genes into 15 clades with grapevine VQ genes, which indicated that at least 15 ancient VQ genes existed before the divergence of the six studied species of Fragaria. Phylogenetic analysis indicated that 28 gene duplication events have occurred in the evolutionary process of the six species of Fragaria. Structural analysis showed that most of the VQ genes have no introns and that VQ proteins in each clade have a similar motif composition. The majority of gene pairs had Ka/Ksratios less than 1, which illustrated that most of the VQ genes underwent purifying selection in the six species of Fragaria. Four types of cis-elements in promoters of VQ genes were detected, which is an important basis for further studies about plant stress responses. Furthermore, the expression analysis of FvVQ genes indicated that these genes are expressed differentially in the examined organs and tissues. The identification of VQ genes and the analysis of VQ gene duplication and polyploidization events in the six species of Fragaria provide important information on the evolutionary fate of VQ genes during the divergence of the six species of Fragaria.

Alternaria blotch, caused by Alternaria alternata f. sp. mali, is a destructive defoliating disease that affects apple worldwide, especially in Asia. Despite previous efforts in characterizing this disease, few have reported on nucleotide binding sites (NBS)-encoding genes in response to pathogen infection. In this study, we characterized the morphological and gene expression response of ‘Golden Delicious’ and ‘Starking’ apples challenged by A. alternata. Inoculation of detached leaves of ‘Golden Delicious’ resulted in a rapid initiation of disease symptoms but a slow spread of the infection from the point of inoculation. In contrast, detached leaves of ‘Starking’ exhibited a delayed visibility of infection but a rapid spread of the infection once it was established. Based upon quantitative reverse transcription polymerase chain reaction data, we characterized the expression of 12 NBS-encoding genes in response to infection by A. alternata. We further characterized the structure, chromosomal location, and...

The Earth has been undergoing climatic changes for centuries, driven by increasing concentration of atmospheric carbon dioxide (CO2). The atmospheric CO2 concentration has been predicted to reach 550–750 μmol mol−1 by 2050, or twice as high as the current level. Much of the research in the last 20–30 years concerning elevated CO2 (eCO2) has been about how plants would respond to the eCO2 at physiological levels. As eCO2 can lead to more frequent drought or extreme high or low temperature, increasingly more research has focused on the interactions between eCO2 and other abiotic stresses. How stresses may affect plant growth and development and productivity, as well as how agricultural practices may be altered to cope with these changes must be determined. These concerns have been the subject of numerous reviews. However, it is only in the last several years that data at the “omics” levels has been available to explore how necessary physiological changes may be brought about in a future complex environment....

With predicted global changes in temperature and precipitation, drought will increasingly impose a challenge to biomass production. Most of the bioenergy crops have some degree of drought susceptibility as revealed for example through measures of low water-use efficiency (WUE). It is imperative to improve drought tolerance and WUE in bioenergy crops for sustainable biomass production in arid and semi-arid regions. Genetics and functional genomics can play critical roles in generating knowledge to inform and aid genetic improvement for drought tolerance in bioenergy crops. The molecular aspects of drought response have been extensively investigated in model plants like Arabidopsis, yet our understanding of the molecular mechanisms underlying drought tolerance in bioenergy crops is limited. Plants in general exhibit various responses to drought stress depending on species and genotype. A rational strategy for studying drought tolerance in bioenergy crops is to translate the knowledge from model plants relat...

Species- or lineage-specific genes can facilitate studying the unique characteristics of biological processes. Updated genome sequences in Populus trichocarpa were screened against thirty newly sequenced or resequenced plant genomes to identify a set of species-specific genes (PtSS). Forty PtSS genes have been isolated with no similarity to any sequence outside the P. trichocarpa genome, therefore have no annotated functions. Protein motif, intron/exon features, subcellular localization and gene expression were analyzed in these PtSS genes. Results reflect their basic genic characters, expression analysis and primary function exploration might provide insight to their possible involvements in lineage specific biological process in woody plants.

Fruit skin coloration is a unique phase in the life cycle of fruiting plants and is mainly attributed to anthocyanin pigments. Anthocyanins are the largest and most diverse group of plant pigments derived from the phenyl propanoid pathway. They are water-soluble phenolic compounds that form part of a large and common group of plant flavonoids. Coloration encompasses several physiological and biochemical changes that happen through differential expression of various developmentally regulated genes. Due to research importance and economic value, Arabidopsis thaliana (chromosome no. = 5) and Vitis vinifera (chromosome no. = 19) have been used for investigations of the structural genes involved in anthocyanin biosynthesis. Thus for this review, V. vinifera is used as a model crop. In anthocyanin biosynthesis, a wide range of constructive genes including phenylalanine ammonia lyase, chalcone synthase and anthocyanidin synthase that are regulated by MYB transcription factors are involved. These genes are coordinately expressed and their levels of expression are positively related to the anthocyanin concentrations. Expression or suppression of the constructive genes contributes to a variety of changes that make fruits visually attractive and edible. Transgenic approaches also have discovered a strong relationship between phenyl propanoid/flavonoid gene expressions for fruit skin coloration. In this study, various developments that have taken place in the last decade with respect to identifying and altering the function of color-related genes are described.

Basic leucine zipper (bZIP) genes are known to play a crucial role in response to various processes in plant as well as abiotic or biotic stress challenges. We have performed an identification and characterization of 50 bZIP genes across the woodland strawberry (Fragaria vesca) genome, which were divided into 10 clades according to the phylogenetic relationship of the strawberry bZIP proteins with those in Arabidopsis and rice. Five categories of intron patterns were observed within basic and hinge regions of the bZIP domains. Some additional conserved motifs have been found with the group specificity. Further, we predicted DNA-binding specificity of the basic and hinge regions as well as dimerization properties of leucine zipper regions, which was consistent with our phylogenetic clade and classified into 20 subfamilies. Across the different developmental stages of 15 organs and two types of fruits, the clade A bZIP members showed different tissue-specific expression patterns and the duplicated genes were differentially regulated, indicating a functional diversification coupled with the expansion of this gene family in strawberry. Under normal growth conditions, mrna11837 and mrna30280 of clade A showed very weak expression levels in organs and fruits, respectively; but higher expression was observed with different set of genes following drought and heat treatment, which may be caused by the separate response pathway between drought and heat treatments.

Apple is one of the most cultivated tree fruits worldwide, and is susceptible to many diseases. Understanding the interactions between the host and pathogen is critical in implementing disease management strategies and developing resistant cultivars. This review provides an update on the interactions of apple with Alternaria alternata apple pathotype, which causes Alternaria blotch, with a brief history about the discovery of the disease and pathogen and its damage and epidemiology. The focus of the review is placed on the physiological and genetic response of the host to pathogen infection, including resistance and susceptibility, and the molecular markers associated with them. Of the response of the pathogen to the host, the emphasis is placed on the role of the selective toxins on pathogenicity and their genetic controls and regulations. The review ends with a perspective on future directions in the research on the apple-A. alternata pathosystem in the era of genomics and post genomics, particularly on...

Genetic engineering of nitrogen assimilation has received considerable attention for its improvement of nitrogen use efficiency in crops. Overexpression of maize Dof1 (ZmDof1), a plant-specific transcription factor, has been reported to promote nitrogen assimilation and growth under low-nitrogen conditions in transgenic Arabidopsis and rice. The present study explored its application in Populus by overexpressing the ZmDof1 gene. Contrary to the results in Arabidopsis and rice, introduction of the ZmDof1 gene did not improve growth of transgenic Populus under different low-nitrogen conditions both in vitro and in greenhouse. Expressions of genes that are involved in nitrogen assimilation and carbon fixation were not induced in ZmDof1-transgenic Populus. Nitrogen/carbon concentrations and photosynthesis rate were not increased in the ZmDof1-transgenic Populus. The results in transgenic Populus overexpressing the ZmDof1 gene indicated that ZmDof1 did not play an important role in nitrogen assimilation under low nitrogen condition in woody Populus plants as it did in annual Arabidopsis and rice plants.

Xyloglucan endotransglycosylase/hydrolases (XTHs) are believed to modify the cell wall structure by cleaving a xyloglucan polymer and transferring the newly generated, potentially reducing, terminal to another xyloglucan. We report here the detailed analysis of 37 Populus trichocarpa XTH genes/proteins in their divergence in both the coding and 5′ promoter regions. Our results show that the Populus XTH genes have experienced whole-genome and local duplications and pre- and post-speciation divergence. Genome-wide and segmental duplications seem to be dominant in subfamily I and III, while tandem duplication seems to be the major mechanism for the subfamily II expansion, which also has higher average ratios of K a/K s compared to those in subfamily I and III. There was a general lack of organ-specific gene expression. In contrast, the expression patterns in subfamily II varied in response to various hormone treatments, with II-A being up-regulated and II-B down-regulated after 2 h of hormone treatment. Expression for this subfamily was verified using the 1.5-kb PtXTH22 promoter that was fused with the GUS reporter gene and transformed into Arabidopsis. The PtXTH22 promoter contains auxin response element, ethylene insensitive 3-like factors, and brassinosteroid response cis-elements. Histochemical GUS staining of transgenic Arabidopsis seedlings confirmed that the PtXTH22 promoter was up-regulated by several hormones.

Species within the genus Populus are among the fastest growing trees in regions with a temperate climate. Not only are they an integral component of ecosystems, but they are also grown commercially for fuel, fiber, and forest products in rural areas of the world. In the late 1970s, they were designated as a bioenergy crop by the U.S. Department of Energy, as a result of research following the oil embargo. Populus species also serve as model trees for plant molecular biology research. In this article, we will review recent progress in the genetic improvement of Populus, considering both classical breeding and genetic engineering for bioenergy, as well as in using transgenics to elucidate gene functionality. A perspective for future improvement of Populus via functional genomics will also be presented.

Alternaria alternata apple pathotype (previously A. mali) causes alternaria blotch disease of apple (Malus ×domestica), which may result in leaf spots and up to 70% premature leaf drop in serious cases. This disease is of worldwide importance but is most serious in eastern Asia (Japan, Korea, and China) and in parts of the United States. The excessive use of fungicides not only adds cost to apple growers, but also pollutes the environment. In this study, we characterized a 5-year F1 population from a cross of a resistant cultivar (Huacui) and a susceptible cultivar (Golden Delicious) consisting of 110 individuals along with 14-year-old parent trees (10 each). A field evaluation of disease severity was conducted in 2008 and 2009 under the natural conditions in Liaoning, China (lat. 40°37′ N, long. 120°44′ E). Based on the field data, 110 F1 plants were divided into five groups. Artificial inoculation was carried out both on the living trees and on the detached leaves in 2009 to ensure that A. alternata apple pathotype was the causative agent. Eighty primer pairs of simple sequence repeat (SSR) were screened against the four genomic DNA pools, respectively, from six highly susceptible F1 plants, six most resistant F1 plants, one tree of the seed parent, and the one tree of the pollen parent. One pair of primers (CH05g07) was shown to be linked to the DNA pools of susceptible F1 and the parent tree, but not to the DNA pools of resistant F1 and parent trees. This primer pair was then used to screen all individual 110 F1 progenies and two parent trees. The differentiation of 103 individuals (97.3%) with the marker matched the field disease resistance rating. This marker was further screened with 20 cultivars with known susceptibility or resistance to A. alternata apple pathotype and its linkage to susceptibility was validated. These results suggest that this marker can be used in marker-assisted selection for resistance/susceptibility to alternaria blotch disease in apple.

Almost all extant plant species have doubled their genomes at least once in their evolutionary histories, resulting in polyploidy which provided a rich genomic resource for evolutionary processes. Moreover, superior polyploid clones have been developed during the process of crop domestication. Polyploid plants generated by evolutionary processes and/or crop domestication have been the intentional or serendipitous focus of research dealing with the dynamics and consequences of genome evolution. One of the new trends in genomics research is to create synthetic polyploid plants which provide materials for studying the initial genomic changes/responses immediately after polyploid formation. Polyploid plants are also used in functional genomics research to study gene expression in a complex genomic background. In this review, we summarize recent progress in genomics research involving ancient, young, and synthetic polyploid plants, with a focus on genome size evolution, genomic diversity, genomic rearrangement, genetic and epigenetic changes in duplicated genes, gene discovery, and comparative genomics. Implications on plant sciences including evolution, functional genomics, and plant breeding are presented. Polyploids will be a focus of genomic research in the future as rapid advances in DNA sequencing technology create unprecedented opportunities for discovering and monitoring genomic and transcriptomic changes. The accumulation of knowledge on polyploid formation, maintenance, and divergence at whole-genome and subgenome levels will not only help plant biologists understand how plants have evolved and diversified, but also assist plant breeders in designing new strategies for crop improvement.

The nucleotide sequence of the entire grapevine genome has been obtained and its draft sequence has served as the model for fruit crops. Many transcription factors involved in stress-resistance pathways have been identified, among which the AP2/ERF is a large, and one of the most important families of transcription factors in plants. In this study, 132 putative transcription factor genes in the AP2/ERF family were identified from the database of the Vitis vinifera (grapevine) genome, and were compared with those from three other completely sequenced genomes, Arabidopsis thaliana, Populus trichocarpa (poplar), and Oryza sativa (rice). The grapevine AP2/ERF genes were classified into four subfamilies, AP2, DREB, ERF, RAV, and a soloist, and there were considerable variations among the subfamily members comparing with those in other species. The amino acid sequence, composition, physical and chemical characterization, phylogenetic tree, conserved domain sequences, function domain, and locations on chromosomes were predicted and analyzed. In silico mapping of the gene loci showed that AP2/ERF family genes of grapevine were localized on 19 linkage groups. The extant AP2/ERF gene family in grapevine apparently resulted from whole genome, segmental and tandem duplications. Several grapevine AP2/ERF genes have been identified which are orthologous to those characterized in Arabidopsis with known functions.

Reliable micropropagation protocols were developed for Japanese honeysuckle (Lonicera japonica Thunb.) and Amur honeysuckle [L. maackii (Rupr.) Maxim], two horticulturally important invasive woody plants. Shoot tips were collected from mature plants and established in vitro on Murashige and Skoog (MS) medium and cultured in various media. Plant growth regulators and basal media had significant effects on proliferation rates. Proliferation of Japanese honeysuckle was highest on Driver and Kuniyuki walnut (DKW) medium with 5 μM (1.1 ppm) 6-benzyladenine (BA), yielding an average of 14 axillary microshoots in 12 weeks, whereas Amur honeysuckle produced 38 axillary microshoots in 12 weeks on MS medium containing 2.5 μM (0.6 ppm) BA with or without 1.25 μM (0.3 ppm) indole-3-butyric acid (IBA). Microcuttings of Japanese honeysuckle were rooted ex vitro with or without treatment with 980 μM (200 ppm) IBA, whereas microcuttings of Amur honeysuckle required treatment with 980 μM (200 ppm) IBA to root. These protocols can be used to continuously produce plant materials for various biotechnological modifications, such as in vitro mutations and genetic engineering for noninvasiveness.

Recent advances in Populus genomics have greatly expanded its popularity as a reference for fundamental as well as applied research in woody plants. In this review, we provide an overview of the state-of-the-science in Populus genomics research, including experimental and computational genomics. We have surveyed and summarized the following: 1) pioneering as well as more recent reports of genetics- and genomics-based investigations in Populus, 2) the positive impact of technological improvements, 3) findings from phylogenetic analyses of gene families, and 4) genomic databases. In the area of Populus experimental genomics, genetic approaches have been advanced to the genome scale with resolution to the gene and/or single nucleotide level. On the other hand, the modern “omics” approaches have been successfully applied to analysis of gene function, such as transcriptome profiling using microarrays as well as the next-generation DNA sequencing technology, and characterization of proteome and metabolome using...

Populus serves as a model tree for biotechnology and molecular biology research due to the availability of the reference genome sequence of Populus trichocarpa (Torr. & Gray) genotype ‘Nisqually-1’. However, ‘Nisqually-1’ has been shown to be very recalcitrant to micropropagation, regeneration and transformation. In this study, a highly efficient micropropagation protocol from greenhouse-grown shoot tips of ‘Nisqually-1’ was established. The optimal micropropagation protocol involves growing in vitro shoots in plant growth regulator-free Murashige and Skoog (MS) basal medium supplemented with 3% sucrose, 0.3% Gelrite® and 5–10 g L−1 of activated charcoal. Plants grown on this medium were significantly longer, and contained significantly higher concentrations of chlorophyll. This highly effective protocol provides a consistent supply of quality leaf and stem materials throughout the year for transformation experiments and other in vitro manipulations, therefore eliminating inconsistency due to seasonal and greenhouse environmental variations and the need for repetitive tissue sterilization.

COBRA proteins have been shown to be involved in both cell wall expansion and/or cellulose deposition. In this paper, we analyzed all 18 COBRA genes (PtCOBRA) from the completely sequenced Populus trichocarpa genome. The 14-member PtCOBRA subfamily I proteins have high similarities to the Arabidopsis (At) COB subfamily, and members with full length sequences were predicted to possess significant potentials for a GPI-anchor site. The 4-member PtCOBRA subfamily II proteins are 45% longer than subfamily I proteins and lack ω-attachment sites at the C terminus, and are more similar to AtCOBL7 subfamily. The expression of the Populus COBRA family genes were regulated in a tissue-specific manner, and were shown to also respond differentially to inductions of hormones and environmental stimuli which affect plant cell expansion. The high levels of expressions, particularly in shoot tip and young root organs, suggests that at least some Populus COBRA genes are likely involved in regulating cell expansion.

Chemical synthesis of DNA sequences provides a powerful tool for modifying genes and for studying gene structure, expression and function. Modified genes and consequently protein/enzymes can bridge genomics and proteomics research or facilitate commercial applications of gene and protein technologies. In this review, we will summarize various strategies, designing softwares and error correction methods for chemical gene synthesis, particularly for the synthesis and assembly of long DNA molecules based on polymerase cycling assembly. Also, we will briefly discuss some of the major applications of chemical synthesis of DNA sequences in basic research and applied areas.

To achieve a thermostable beta-glucuronidase (GUS) and identify key mutation sites, we applied in vitro directed evolution strategy through DNA shuffling and obtained a highly thermostable mutant GUS gene, gus-tr, after four rounds of DNA shuffling and screening. This variant had mutations in 15 nucleic acid sites, resulting in changes in 12 amino acids (AAs). Using gus-tr as the template, we further performed site-directed mutagenesis to reverse the individual mutation to the wild-type protein. We found that six sites (Q493R, T509A, M532T, N550S, G559S and N566S) present in GUS-TR3337, were the key AAs needed to confer its high thermostability. Of these, Q493R and T509A were not reported previously as important residues for thermostability of GUS. Furthermore, all of these six mutations must be present concurrently to confer the high thermostability. We expressed the gus-tr3337 gene and purified the GUS-TR3337 protein that contained the six AA mutations. Compared with the wild-type protein which lost its activity completely after 10 min at 70 degrees C, the mutant GUS-TR3337 protein retained 75% of its activity when heated at 80 degrees C for 10 min. The GUS-TR3337 exhibited high activity even heated at 100 degrees C for 30 min on nitrocellulose filter. The comparison of molecular models of the mutated and wild-type enzyme revealed the relation of protein function and these structural modifications.

Background Genetic engineering of plants that results in successful establishment of new biochemical or regulatory pathways requires stable introduction of one or more genes into the plant genome. It might also be necessary to down-regulate or turn off expression of endogenous genes in order to reduce activity of competing pathways. An established way to knockdown gene expression in plants is expressing a hairpin-RNAi construct, eventually leading to degradation of a specifically targeted mRNA. Knockdown of multiple genes that do not share homologous sequences is still challenging and involves either sophisticated cloning strategies to create vectors with different serial expression constructs or multiple transformation events that is often restricted by a lack of available transformation markers. Results Synthetic RNAi fragments were assembled in yeast carrying homologous sequences to six or seven non-family genes and introduced into pAGRIKOLA. Transformation of Arabidopsis thaliana and subsequent expression analysis of targeted genes proved efficient knockdown of all target genes. Conclusions We present a simple and cost-effective method to create constructs to simultaneously knockdown multiple non-family genes or genes that do not share sequence homology. The presented method can be applied in plant and animal synthetic biology as well as traditional plant and animal genetic engineering. Electronic supplementary material The online version of this article (doi:10.1186/s13007-016-0116-8) contains supplementary material, which is available to authorized users.

The mitotic inhibitors, colchicine and oryzalin, were evaluated for their effects on callus, adventitious shoot formation, and tetraploid induction of Euphorbia pulchurrima `Winter Rose'. In vitro grown leaf sections were placed on various media supplemented with either colchicine or oryzalin at various concentrations for 1 to 4 days. Colchicine was less damaging to leaf tissues than oryzalin. On various colchicine-containing media, prolific calluses were produced and adventitious shoot formation was observed. Regenerated shoots were found to be diploid as determined by flow cytometry. On media supplemented with oryzalin (28.9 μm to 144 μm), leaf tissues produced callus but failed to form adventitious shoots. Samples of calluses produced on oryzalin-containing media were subject to analysis using flow cytometry and were found to be diploid. These results suggest that the colchicine is less toxic on poinsettia tissues and shoot induction than oryzalin. Additional experiments are needed to establish a protocol for in vitro induction of poinsettia tetraploid with colchicine and oryzalin.

Invasive plants, one of the most devastating ecological problems in the 21st century, cause an estimated $35 billion loss per year to the economy in the United States alone. More than 50% of all invasive plant species and 85% of invasive woody plant species were introduced originally for ornamental and landscape use. Because many non-native ornamentals are commercially important and widely utilized for various purposes, completely banning their use and prohibiting their import are unpractical solutions. On the other hand, currently used methods to control the spread of non-native plants are ineffective, expensive, or environmentally problematic. Recent advances in plant molecular biology and plant genetic transformation may enable us to create sterile cultivars of these non-native ornamental crops of high commercial value. The use of sterile cultivars should reduce or eliminate the undesirable spread of some non-native invasive plants into natural areas.

Vegetative propagation by cuttings is critical to the horticultural industry, particularly the ornamental and fruit sections. This article briefly reviews the traditional approaches to improve root formation, specifically with hardwood cuttings. The focus of this review has been placed on the nontraditional strategies, including infection with Agrobacterium rhizogenes and genetic engineering with rol genes, or ro/B gene alone from A. rhizogenes and iaaM gene from auxin biosynthesis genes (iaaM, iaaH) from A. tumefaciens. Recent results with these new strategies have shown great potential in enhancing adventitious root formation in difficult-to-root woody species. The future perspective in understanding adventitious rooting mechanisms and new strategies for improving rooting are also discussed.

It is widely accepted that gene duplication is a primary source of genetic novelty. However, the evolutionary fate of duplicated genes remains largely unresolved. The classical Ohno's Duplication-Retention-Non/Neofunctionalization theory, and the recently proposed alternatives such as subfunctionalization or duplication-degeneration-complementation, and subneofunctionalization, each can explain one or more aspects of gene fate after duplication. Duplicated genes are also affected by epigenetic changes. We constructed a phylogenetic tree using Dof (DNA binding with one finger) protein sequences from poplar (Populus trichocarpa) Torr. & Gray ex Brayshaw, Arabidopsis (Arabidopsis thaliana), and rice (Oryza sativa). From the phylogenetic tree, we identified 27 pairs of paralogous Dof genes in the terminal nodes. Analysis of protein motif structure of the Dof paralogs and their ancestors revealed six different gene fates after gene duplication. Differential protein methylation was revealed between a pair of duplicated poplar Dof genes, which have identical motif structure and similar expression pattern, indicating that epigenetics is involved in evolution. Analysis of reverse transcription-PCR, massively parallel signature sequencing, and microarray data revealed that the paralogs differ in expression pattern. Furthermore, analysis of nonsynonymous and synonymous substitution rates indicated that divergence of the duplicated genes was driven by positive selection. About one-half of the motifs in Dof proteins were shared by non-Dof proteins in the three plants species, indicating that motif co-option may be one of the forces driving gene diversification. We provided evidence that the Ohno's Duplication-Retention-Non/Neofunctionalization, subfunctionalization/duplication-degeneration-complementation, and subneofunctionalization hypotheses are complementary with, not alternative to, each other.

Mannans are widespread hemicellulosic polysaccharides in plant cell walls. Hydrolysis of the internal beta-1,4-D: -mannopyranosyl linkage in the backbone of mannans is catalyzed by endo-beta-mannanase. Plant endo-beta-mannanase has been well studied for its function in seed germination. Its involvement in other plant biological processes, however, remains poorly characterized or elusive. The completed genome sequences of Arabidopsis (Arabidopsis thaliana), rice (Oryza sativa), and poplar (Populus trichocarpa) provide an opportunity to conduct comparative genomic analysis of endo-beta-mannanase genes in these three species. In silico sequence analysis led to the identification of eight, nine and 11 endo-beta-mannanase genes in the genomes of Arabidopsis, rice, and poplar, respectively. Sequence comparisons revealed the conserved amino acids and motifs that are critical for the active site of endo-beta-mannanases. Intron/exon structure analysis in conjunction with phylogenetic analysis implied that both intron gain and intron loss has played roles in the evolution of endo-beta-mannanase genes. The phylogenetic analysis that included the endo-beta-mannanases from plants and other organisms implied that plant endo-beta-mannanases have an ancient evolutionary origin. Comprehensive expression analysis of all Arabidopsis and rice endo-beta-mannanase genes showed divergent expression patterns of individual genes, suggesting that the enzymes encoded by these genes, while carrying out the same biochemical reaction, are involved in diverse biological processes.