The following list is a collection of papers on transcriptomes of members of the Brassicaceae. Reviews and proceedings abstracts have been omitted. Results are shown in reverse chronological order.
White, O. W., B. Doo, et al. (2016) TRANSCRIPTOME SEQUENCING AND SIMPLE SEQUENCE REPEAT MARKER DEVELOPMENT FOR THREE MACARONESIAN ENDEMIC PLANT SPECIES Applications in Plant Sciences 4: 1600050
10.3732/apps.1600050
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Premise of the study: Oceanic islands offer unparalleled opportunities to investigate evolutionary processes such as adaptation and speciation. However, few genomic resources are available for oceanic island endemics. In this study, we publish transcriptome sequences from three Macaronesian endemic plant species (Argyranthemum broussonetii [Asteraceae], Descurainia bourgaeana [Brassicaceae], and Echium wildpretii [Boraginaceae]) that are representative of lineages that have radiated in the region. In addition, the utility of transcriptome data for marker development is demonstrated. Methods and Results: Transcriptomes from the three plant species were sequenced, assembled, and annotated. Between 1972 and 2282 simple sequence repeats (SSRs) were identified for each taxon. Primers were designed and tested for 30 of the candidate SSRs identified in Argyranthemum, of which 12 amplified well across three species and eight were polymorphic. Conclusions: We demonstrate here that a single transcriptome sequence is sufficient to identify hundreds of polymorphic SSR markers. The SSRs are applicable to a wide range of questions relating to the evolution of island lineages.
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Voutsina, N., A. C. Payne, et al. (2016) Characterization of the watercress (Nasturtium officinale R. Br.; Brassicaceae) transcriptome using RNASeq and identification of candidate genes for important phytonutrient traits linked to human health Bmc Genomics 17: 378
10.1186/s12864-016-2704-4
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Background: Consuming watercress is thought to provide health benefits as a consequence of its phytonutrient composition. However, for watercress there are currently limited genetic resources underpinning breeding efforts for either yield or phytonutritional traits. In this paper, we use RNASeq data from twelve watercress accessions to characterize the transcriptome, perform candidate gene mining and conduct differential expression analysis for two key phytonutritional traits: antioxidant (AO) capacity and glucosinolate (GLS) content. Results: The watercress transcriptome was assembled to 80,800 transcripts (48,732 unigenes); 71 % of which were annotated based on orthology to Arabidopsis. Differential expression analysis comparing watercress accessions with ‘high’ and ‘low’ AO and GLS resulted in 145 and 94 differentially expressed loci for AO capacity and GLS respectively. Differentially expressed loci between high and low AO watercress were significantly enriched for genes involved in plant defence and response to stimuli, in line with the observation that AO are involved in plant stress-response. Differential expression between the high and low GLS watercress identified links to GLS regulation and also novel transcripts warranting further investigation. Additionally, we successfully identified watercress orthologs for Arabidopsis phenylpropanoid, GLS and shikimate biosynthesis pathway genes, and compiled a catalogue of polymorphic markers for future applications. Conclusions: Our work describes the first transcriptome of watercress and establishes the foundation for further molecular study by providing valuable resources, including sequence data, annotated transcripts, candidate genes and markers.
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Qiao, Q., Q. Wang, et al. (2016) Transcriptome sequencing of Crucihimalaya himalaica (Brassicaceae) reveals how Arabidopsis close relative adapt to the Qinghai-Tibet Plateau Scientific Reports 6: 21729
10.1038/srep21729
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The extreme environment of the Qinghai-Tibet Plateau (QTP) provides an ideal natural laboratory for studies on adaptive evolution. Few genome/transcriptome based studies have been conducted on how plants adapt to the environments of QTP compared to numerous studies on vertebrates. Crucihimalaya himalaica is a close relative of Arabidopsis with typical QTP distribution, and is hoped to be a new model system to study speciation and ecological adaptation in extreme environment. In this study, we de novo generated a transcriptome sequence of C. himalaica, with a total of 49,438 unigenes. Compared to five relatives, 10,487 orthogroups were shared by all six species, and 4,286 orthogroups contain putative single copy gene. Further analysis identified 487 extremely significantly positively selected genes (PSGs) in C. himalaica transcriptome. Theses PSGs were enriched in functions related to specific adaptation traits, such as response to radiation, DNA repair, nitrogen metabolism, and stabilization of membrane. These functions are responsible for the adaptation of C. himalaica to the high radiation, soil depletion and low temperature environments on QTP. Our findings indicate that C. himalaica has evolved complex strategies for adapting to the extreme environments on QTP and provide novel insights into genetic mechanisms of highland adaptation in plants.
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Lee, Y. P., C. Funk, et al. (2016) Salt stress responses in a geographically diverse collection of Eutrema/Thellungiella spp. accessions Functional Plant Biology 43: 590-606. 10.1071/fp15285
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Salinity strongly impairs plant growth and development. Natural genetic variation can be used to dissect complex traits such as plant salt tolerance. We used 16 accessions of the halophytic species Eutrema salsugineum (previously called Thellungiella salsuginea (Pallas) O.E. Schulz, Thellungiella halophila (C.A. Meyer) O.E. Schulz and Thellungiella botschantzevii D. A. German to investigate their natural variation in salinity tolerance. Although all accessions showed survival and growth up to 700mM NaCl in hydroponic culture, their relative salt tolerance varied considerably. All accessions accumulated the compatible solutes proline, sucrose, glucose and fructose and the polyamines putrescine and spermine. Relative salt tolerance was not correlated with the content of any of the investigated solutes. We compared the metabolomes and transcriptomes of Arabidopsis thaliana (L. Heynh.) Col-0 and E. salsugineum Yukon under control and salt stress conditions. Higher content of several metabolites in Yukon compared with Col-0 under control conditions indicated metabolic pre-adaptation to salinity in the halophyte. Most metabolic salt responses in Yukon took place at 200mM NaCl, whereas few additional changes were observed between 200 and 500 mM. The opposite trend was observed for the transcriptome, with only little overlap between salt-regulated genes in the two species. In addition, only about half of the salt-regulated Yukon unigenes had orthologues in Col-0.
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Geiser, C., T. Mandakova, et al. (2016) Repeated Whole-Genome Duplication, Karyotype Reshuffling, and Biased Retention of Stress-Responding Genes in Buckler Mustard Plant Cell 28: 17-27. 10.1105/tpc.15.00791
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Whole-genome duplication (WGD) is usually followed by gene loss and karyotype repatterning. Despite evidence of new adaptive traits associated with WGD, the underpinnings and evolutionary significance of such genome fractionation remain elusive. Here, we use Buckler mustard (Biscutella laevigata) to infer processes that have driven the retention of duplicated genes after recurrent WGDs. In addition to the beta- and alpha-WGD events shared by all Brassicaceae, cytogenetic and transcriptome analyses revealed two younger WGD events that occurred at times of environmental changes in the clade of Buckler mustard (Biscutelleae): a mesopolyploidy event from the late Miocene that was followed by considerable karyotype reshuffling and chromosome number reduction and a neopolyploidy event during the Pleistocene. Although a considerable number of the older duplicates presented signatures of retention under positive selection, the majority of retained duplicates arising from the younger mesopolyploidy WGD event matched predictions of the gene balance hypothesis and showed evidence of strong purifying selection as well as enrichment in gene categories responding to abiotic stressors. Retention of large stretches of chromosomes for both genomic copies supported the hypothesis that cycles of WGD and biased fractionation shaped the genome of this stress-tolerant polypolyloid, promoting the adaptive recruitment of stress-responding genes in the face of environmental challenges.
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Auguy, F., M. Fahr, et al. (2016) Transcriptome Changes in Hirschfeldia incana in Response to Lead Exposure Frontiers in Plant Science 6: 1231
10.3389/fpls.2015.01231
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Hirschfeldia incana, a pseudometallophyte belonging to the Brassicaceae family and widespread in the Mediterranean region, was selected for its ability to grow on soils contaminated by lead (Pb). The global comparison of gene expression using microarrays between a plant susceptible to Pb (Arabidopsis thaliana) and a Pb tolerant plant (H. incana) enabled the identification of a set of specific genes expressed in response to lead exposure. Three groups of genes were particularly over-represented by the Pb exposure in the biological processes categorized as photosynthesis, cell wall, and metal handling. Each of these gene groups was shown to be directly involved in tolerance or in protection mechanisms to the phytotoxicity associated with Pb. Among these genes, we demonstrated that MT2b, a metallothionein gene, was involved in lead accumulation, confirming the important role of metallothioneins in the accumulation and the distribution of Pb in leaves. On the other hand, several genes involved in biosynthesis of ABA were shown to be up regulated in the roots and shoots of H. incana treated with Pb, suggesting that ABA-mediated signaling is a possible mechanism in response to Pb treatment in H. incana. This latest finding is an important research direction for future studies.
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Stockenhuber, R., S. Zoller, et al. (2015) Efficient Detection of Novel Nuclear Markers for Brassicaceae by Transcriptome Sequencing Plos One 10: e0128181
10.1371/journal.pone.0128181
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The lack of DNA sequence information for most non-model organisms impairs the design of primers that are universally applicable for the study of molecular polymorphisms in nuclear markers. Next-generation sequencing (NGS) techniques nowadays provide a powerful approach to overcome this limitation. We present a flexible and inexpensive method to identify large numbers of nuclear primer pairs that amplify in most Brassicaceae species. We first obtained and mapped NGS transcriptome sequencing reads from two of the distantly related Brassicaceae species, Cardamine hirsuta and Arabis alpina, onto the Arabidopsis thaliana reference genome, and then identified short conserved sequence motifs among the three species bioinformatically. From these, primer pairs to amplify coding regions (nuclear protein coding loci, NPCL) and exon-primed intron-crossing sequences (EPIC) were developed. We identified 2,334 universally applicable primer pairs, targeting 1,164 genes, which provide a large pool of markers as readily usable genomic resource that will help addressing novel questions in the Brassicaceae family. Testing a subset of the newly designed nuclear primer pairs revealed that a great majority yielded a single amplicon in all of the 30 investigated Brassicaceae taxa. Sequence analysis and phylogenetic reconstruction with a subset of these markers on different levels of phylogenetic divergence in the mustard family were compared with previous studies. The results corroborate the usefulness of the newly developed primer pairs, e.g., for phylogenetic analyses or population genetic studies. Thus, our method provides a cost-effective approach for designing nuclear loci across a broad range of taxa and is compatible with current NGS technologies.
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Mucha, S., D. Walther, et al. (2015) Substantial reprogramming of the Eutrema salsugineum (Thellungiella salsuginea) transcriptome in response to UV and silver nitrate challenge Bmc Plant Biology 15: 137
10.1186/s12870-015-0506-5
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Background: Cruciferous plants synthesize a large variety of tryptophan-derived phytoalexins in response to pathogen infection, UV irradiation, or high dosages of heavy metals. The major phytoalexins of Eutrema salsugineum (Thellungiella salsuginea), which has recently been established as an extremophile model plant, are probably derivatives of indole glucosinolates, in contrast to Arabidopsis, which synthesizes characteristic camalexin from the glucosinolate precursor indole-3-acetaldoxime. Results: The transcriptional response of E. salsugineum to UV irradiation and AgNO3 was monitored by RNAseq and microarray analysis. Most transcripts (respectively 70% and 78%) were significantly differentially regulated and a large overlap between the two treatments was observed (54% of total). While core genes of the biosynthesis of aliphatic glucosinolates were repressed, tryptophan and indole glucosinolate biosynthetic genes, as well as defence-related WRKY transcription factors, were consistently upregulated. The putative Eutrema WRKY33 ortholog was functionally tested and shown to complement camalexin deficiency in Atwrky33 mutant. Conclusions: In E. salsugineum, UV irradiation or heavy metal application resulted in substantial transcriptional reprogramming. Consistently induced genes of indole glucosinolate biosynthesis and modification will serve as candidate genes for the biosynthesis of Eutrema-specific phytoalexins.
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Rohini, G., V. Mohit, et al. (2014) Deep Transcriptome Sequencing of Wild Halophyte Rice, Porteresia coarctata, Provides Novel Insights into the Salinity and Submergence Tolerance Factors DNA Research 21: 69-84. 10.1093/dnares/dst042
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Porteresia coarctata is a wild relative of rice with capability of high salinity and submergence tolerance. The transcriptome analyses of Porteresia can lead to the identification of candidate genes involved in salinity and submergence tolerance. We sequenced the transcriptome of Porteresia under different conditions using Illumina platform and generated about 375 million high-quality reads. After optimized assembly, a total of 152 367 unique transcript sequences with average length of 794 bp were obtained. Many of these sequences might represent fragmented transcripts. Functional annotation revealed the presence of genes involved in diverse cellular processes and 2749 transcription factor (TF)-encoding genes in Porteresia. The differential gene expression analyses identified a total of 15 158 genes involved in salinity and/or submergence response(s). The stress-responsive members of different TF families, including MYB, bHLH, AP2-EREBP, WRKY, bZIP and NAC, were identified. We also revealed key metabolic pathways, including amino acid biosynthesis, hormone biosynthesis, secondary metabolite biosynthesis, carbohydrate metabolism and cell wall structures, involved in stress tolerance in Porteresia. The transcriptome analyses of Porteresia are expected to highlight genes/pathways involved in salinity and submergence tolerance of this halophyte species. The data can serve as a resource for unravelling the underlying mechanism and devising strategies to engineer salinity and submergence tolerance in rice.
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Oh, D. H., H. Hong, et al. (2014) Genome Structures and Transcriptomes Signify Niche Adaptation for the Multiple-Ion-Tolerant Extremophyte Schrenkiella parvula Plant Physiology 164: 2123-2138. 10.1104/pp.113.233551
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Schrenkiella parvula (formerly Thellungiella parvula), a close relative of Arabidopsis (Arabidopsis thaliana) and Brassica crop species, thrives on the shores of Lake Tuz, Turkey, where soils accumulate high concentrations of multiple-ion salts. Despite the stark differences in adaptations to extreme salt stresses, the genomes of S. parvula and Arabidopsis show extensive synteny. S. parvula completes its life cycle in the presence of Na+, K+, Mg2(+), Li+, and borate at soil concentrations lethal to Arabidopsis. Genome structural variations, including tandem duplications and translocations of genes, interrupt the colinearity observed throughout the S. parvula and Arabidopsis genomes. Structural variations distinguish homologous gene pairs characterized by divergent promoter sequences and basal-level expression strengths. Comparative RNA sequencing reveals the enrichment of ion-transport functions among genes with higher expression in S. parvula, while pathogen defense-related genes show higher expression in Arabidopsis. Key stress-related ion transporter genes in S. parvula showed increased copy number, higher transcript dosage, and evidence for subfunctionalization. This extremophyte offers a framework to identify the requisite adjustments of genomic architecture and expression control for a set of genes found in most plants in a way to support distinct niche adaptation and lifestyles.
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Milner, M. J., N. Mitani-Ueno, et al. (2014) Root and shoot transcriptome analysis of two ecotypes of Noccaea caerulescens uncovers the role of NcNramp1 in Cd hyperaccumulation Plant Journal 78: 398-410. 10.1111/tpj.12480
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The Zn/Cd hyperaccumulator, Noccaea caerulescens, has been studied extensively for its ability to accumulate high levels of Zn and Cd in its leaves. Previous studies have indicated that the Zn and Cd hyperaccumulation trait exhibited by this species involves different transport and tolerance mechanisms. It has also been well documented that certain ecotypes of N.caerulescens are much better Cd hyperaccumulators than others. However, there does not seem to be much ecotypic variation for Zn hyperaccumulation in N.caerulescens. In this study we employed a comparative transcriptomics approach to look at root and shoot gene expression in Ganges and Prayon plants in response to Cd stress to identify transporter genes that were more highly expressed in either the roots or shoots of the superior Cd accumulator, Ganges. Comparison of the transcriptomes from the two ecotypes of Noccaea caerulescens identified a number of genes that encoded metal transporters that were more highly expressed in the Ganges ecotype in response to Cd stress. Characterization of one of these transporters, NcNramp1, showed that it is involved in the influx of Cd across the endodermal plasma membrane and thus may play a key role in Cd flux into the stele and root-to-shoot Cd transport. NcNramp1 may be one of the main transporters involved in Cd hyperaccumulation in N.caerulescens and copy number variation appears to be the main reason for high NcNramp1 gene expression underlying the increased Cd accumulation in the Ganges ecotype.
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Lin, Y. F., E. I. Severing, et al. (2014) A comprehensive set of transcript sequences of the heavy metal hyperaccumulator Noccaea caerulescens Frontiers in Plant Science 5: 261
10.3389/fpls.2014.00261
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Noccaea caerulescens is an extremophile plant species belonging to the Brassicaceae family. It has adapted to grow on soils containing high, normally toxic, concentrations of metals such as nickel, zinc, and cadmium. Next to being extremely tolerant to these metals, it is one of the few species known to hyperaccumulate these metals to extremely high concentrations in their aboveground biomass. In order to provide additional molecular resources for this model metal hyperaccumulator species to study and understand the mechanism of adaptation to heavy metal exposure, we aimed to provide a comprehensive database of transcript sequences for N. caerulescens. In this study, 23,830 transcript sequences (isotigs) with an average length of 1025 bp were determined for roots, shoots and inflorescences of N. caerulescens accession “Ganges” by Roche GS-FLEX 454 pyrosequencing. These isotigs were grouped into 20,378 isogroups, representing potential genes. This is a large expansion of the existing N. caerulescens transcriptome set consisting of 3705 unigenes. When translated and compared to a Brassicaceae proteome set, 22,232 (93.2%) of the N. caerulescens isotigs (corresponding to 19,191 isogroups) had a significant match and could be annotated accordingly. Of the remaining sequences, 98 isotigs resembled non-plant sequences and 1386 had no significant similarity to any sequence in the GenBank database. Among the annotated set there were many isotigs with similarity to metal homeostasis genes or genes for glucosinolate biosynthesis. Only for transcripts similar to Metallothionein3 (MT3), clear evidence for an additional copy was found. This comprehensive set of transcripts is expected to further contribute to the discovery of mechanisms used by N. caerulescens to adapt to heavy metal exposure.
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Kagale, S., S. J. Robinson, et al. (2014) Polyploid Evolution of the Brassicaceae during the Cenozoic Era Plant Cell 26: 2777-2791. 10.1105/tpc.114.126391
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The Brassicaceae (Cruciferae) family, owing to its remarkable species, genetic, and physiological diversity as well as its significant economic potential, has become a model for polyploidy and evolutionary studies. Utilizing extensive transcriptome pyrosequencing of diverse taxa, we established a resolved phylogeny of a subset of crucifer species. We elucidated the frequency, age, and phylogenetic position of polyploidy and lineage separation events that have marked the evolutionary history of the Brassicaceae. Besides the well-known ancient a (47 million years ago [Mya]) and beta (124 Mya) paleopolyploidy events, several species were shown to have undergone a further more recent (similar to 7 to 12 Mya) round of genome multiplication. We identified eight whole-genome duplications corresponding to at least five independent neo/mesopolyploidy events. Although the Brassicaceae family evolved from other eudicots at the beginning of the Cenozoic era of the Earth (60 Mya), major diversification occurred only during the Neogene period (0 to 23 Mya). Remarkably, the widespread species divergence, major polyploidy, and lineage separation events during Brassicaceae evolution are clustered in time around epoch transitions characterized by prolonged unstable climatic conditions. The synchronized diversification of Brassicaceae species suggests that polyploid events may have conferred higher adaptability and increased tolerance toward the drastically changing global environment, thus facilitating species radiation.
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Halimaa, P., D. Blande, et al. (2014) Comparative transcriptome analysis of the metal hyperaccumulator Noccaea caerulescens Frontiers in Plant Science 5: 213
10.3389/fpls.2014.00213
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The metal hyperaccumulator Noccaea caerulescens is an established model to study the adaptation of plants to metalliferous soils. Various comparators have been used in these studies. The choice of suitable comparators is important and depends on the hypothesis to be tested and methods to be used. In high-throughput analyses such as microarray, N. caerulescens has been compared to non-tolerant, non-accumulator plants like Arabidopsis thaliana or Thlaspi arvense rather than to the related hypertolerant or hyperaccumulator plants. An underutilized source is N. caerulescens populations with considerable variation in their capacity to accumulate and tolerate metals. Whole transcriptome sequencing (RNA-Seq) is revealing interesting variation in their gene expression profiles. Combining physiological characteristics of N. caerulescens accessions with their RNA-Seq has a great potential to provide detailed insight into the underlying molecular mechanisms, including entirely new gene products. In this review we will critically consider comparative transcriptome analyses carried out to explore metal hyperaccumulation and hypertolerance of N. caerulescens, and demonstrate the potential of RNA-Seq analysis as a tool in evolutionary genomics.
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Verbruggen, N., M. Hanikenne, et al. (2013) A more complete picture of metal hyperaccumulation through next-generation sequencing technologies Frontiers in Plant Science 4: 388
10.3389/fpls.2013.00388
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The mechanistic understanding of metal hyperaccumulation has benefitted immensely from the use of molecular genetics tools developed for Arabidopsis thaliana. The revolution in DNA sequencing will enable even greater strides in the near future, this time not restricted to the family Brassicaceae. Reference genomes are within reach for many ecologically interesting species including heterozygous outbreeders. They will allow deep RNA-seq transcriptome studies and the re-sequencing of contrasting individuals to unravel the genetic basis of phenotypic variation. Cell-type specific transcriptome analyses, which will be essential for the dissection of metal translocation pathways in hyperaccumulators, can be achieved through the combination of RNA-seq and translatome approaches. Affordable high-resolution genotyping of many individuals enables the elucidation of quantitative trait loci in intra- and interspecific crosses as well as through genome-wide association mapping across large panels of accessions. Furthermore, genome-wide scans have the power to detect loci under recent selection. Together these approaches will lead to a detailed understanding of the evolutionary path towards the emergence of hyperaccumulation traits.
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Physiol, M. P. I. M. P. (2013) Eutrema salsugineum Transcriptome or Gene expression European Nucleotide Archive [toggle]
Thellungiella salsuginea has attracted increasing interest as an alternative to Arabidopsis as a plant model species that possesses high tolerance of various abiotic stresses. ESTs were generated from a non-normalized and a normalized library synthesized from RNA pooled from plant samples from different developmental stages and grown under abiotic stress conditions.
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Minzu University of, C. (2013) Eutrema halophilum Transcriptome or Gene expression European Nucleotide Archive [toggle]
As a salt tolerant relative of Arabidopsis, Thellungiella salsuginea has increasingly been used in studies of plant salinity tolerance. In the past years, large-scale screening of salt stress-related gene was hindered by lack of sufficient transcriptomic and genomic data in public databases. Thus, high throughput transcriptome sequencing and subsequent digital gene expression analysis is helpful to generate a large amount of transcript sequences for gene discovery and perform high-throughput profiling of high salinity regulated genes in Thellungiella.
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Lee, Y., F. M. Giorgi, et al. (2013) Transcriptome sequencing and microarray design for functional genomics in the extremophile Arabidopsis relative Thellungiella salsuginea ( Eutrema salsugineum) Bmc Genomics 14: (14 November 2013)-(2014 November 2013). [toggle]
Background: Most molecular studies of plant stress tolerance have been performed with Arabidopsis thaliana, although it is not particularly stress tolerant and may lack protective mechanisms required to survive extreme environmental conditions. Thellungiella salsuginea has attracted interest as an alternative plant model species with high tolerance of various abiotic stresses. While the T. salsuginea genome has recently been sequenced, its annotation is still incomplete and transcriptomic information is scarce. In addition, functional genomics investigations in this species are severely hampered by a lack of affordable tools for genome-wide gene expression studies. Results: Here, we report the results of Thellungiella de novo transcriptome assembly and annotation based on 454 pyrosequencing and development and validation of a T. salsuginea microarray. ESTs were generated from a non-normalized and a normalized library synthesized from RNA pooled from samples covering different tissues and abiotic stress conditions. Both libraries yielded partially unique sequences, indicating their necessity to obtain comprehensive transcriptome coverage. More than 1 million sequence reads were assembled into 42,810 unigenes, approximately 50% of which could be functionally annotated. These unigenes were compared to all available Thellungiella genome sequence information. In addition, the groups of Late Embryogenesis Abundant (LEA) proteins, Mitogen Activated Protein (MAP) kinases and protein phosphatases were annotated in detail. We also predicted the target genes for 384 putative miRNAs. From the sequence information, we constructed a 44 k Agilent oligonucleotide microarray. Comparison of same-species and cross-species hybridization results showed superior performance of the newly designed array for T. salsuginea samples. The developed microarrays were used to investigate transcriptional responses of T. salsuginea and Arabidopsis during cold acclimation using the MapMan software. Conclusions: This study provides the first comprehensive transcriptome information for the extremophile Arabidopsis relative T. salsuginea. The data constitute a more than three-fold increase in the number of publicly available unigene sequences and will greatly facilitate genome annotation. In addition, we have designed and validated the first genome-wide microarray for T. salsuginea, which will be commercially available. Together with the publicly available MapMan software this will become an important tool for functional genomics of plant stress tolerance.
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Champigny, M. J., W. W. L. Sung, et al. (2013) RNA-Seq effectively monitors gene expression in Eutrema salsugineum plants growing in an extreme natural habitat and in controlled growth cabinet conditions Bmc Genomics 14: 578
10.1186/1471-2164-14-578
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Background: The investigation of extremophile plant species growing in their natural environment offers certain advantages, chiefly that plants adapted to severe habitats have a repertoire of stress tolerance genes that are regulated to maximize plant performance under physiologically challenging conditions. Accordingly, transcriptome sequencing offers a powerful approach to address questions concerning the influence of natural habitat on the physiology of an organism. We used RNA sequencing of Eutrema salsugineum, an extremophile relative of Arabidopsis thaliana, to investigate the extent to which genetic variation and controlled versus natural environments contribute to differences between transcript profiles. Results: Using 10 million cDNA reads, we compared transcriptomes from two natural Eutrema accessions (originating from Yukon Territory, Canada and Shandong Province, China) grown under controlled conditions in cabinets and those from Yukon plants collected at a Yukon field site. We assessed the genetic heterogeneity between individuals using single-nucleotide polymorphisms (SNPs) and the expression patterns of 27,016 genes. Over 39,000 SNPs distinguish the Yukon from the Shandong accessions but only 4,475 SNPs differentiated transcriptomes of Yukon field plants from an inbred Yukon line. We found 2,989 genes that were differentially expressed between the three sample groups and multivariate statistical analyses showed that transcriptomes of individual plants from a Yukon field site were as reproducible as those from inbred plants grown under controlled conditions. Predicted functions based upon gene ontology classifications show that the transcriptomes of field plants were enriched by the differential expression of light- and stress-related genes, an observation consistent with the habitat where the plants were found. Conclusion: Our expectation that comparative RNA-Seq analysis of transcriptomes from plants originating in natural habitats would be confounded by uncontrolled genetic and environmental factors was not borne out. Moreover, the transcriptome data shows little genetic variation between laboratory Yukon Eutrema plants and those found at a field site. Transcriptomes were reproducible and biological associations meaningful whether plants were grown in cabinets or found in the field. Thus RNA-Seq is a valuable approach to study native plants in natural environments and this technology can be exploited to discover new gene targets for improved crop performance under adverse conditions.
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Voelckel, C., N. Gruenheit, et al. (2012) Chips and tags suggest plant-environment interactions differ for two alpine Pachycladon species Bmc Genomics 13: 322
10.1186/1471-2164-13-322
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Background: Expression profiling has been proposed as a means for screening non-model organisms in their natural environments to identify genes potentially important in adaptive diversification. Tag profiling using high throughput sequencing is a relatively low cost means of expression profiling with deep coverage. However the extent to which very short cDNA sequences can be effectively used in screening for candidate genes is unclear. Here we investigate this question using an evolutionarily distant as well as a closely related transcriptome for referencing tags. We do this by comparing differentially expressed genes and processes between two closely related allopolyploid species of Pachycladon which have distinct altitudinal preferences in the New Zealand Southern Alps. We validate biological inferences against earlier microarray analyses. Results: Statistical and gene annotation enrichment analyses of tag profiles identified more differentially expressed genes of potential adaptive significance than previous analyses of array-based expression profiles. These include genes involved in glucosinolate metabolism, flowering time, and response to cold, desiccation, fungi and oxidation. In addition, despite the short length of 20mer tags, we were able to infer patterns of homeologous gene expression for 700 genes in our reference library of 7,128 full-length Pachycladon ESTs. We also demonstrate that there is significant information loss when mapping tags to the non-conspecific reference transcriptome of A. thaliana as opposed to P. fastigiatum ESTs but also describe mapping strategies by which the larger collection of A. thaliana ESTs can be used as a reference. Conclusion: When coupled with a reference transcriptome generated using RNA-seq, tag sequencing offers a promising approach for screening natural populations and identifying candidate genes of potential adaptive significance. We identify computational issues important for the successful application of tag profiling in a non-model allopolyploid plant species.
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Renou, J.-P. P., L. Soubigou-Taconnat, et al. (2012) Osmotic stress tolerance in Thellungiella halophila ArrayExpress Archive [toggle]
Suitability of CATMA for the analysis of the transcriptome of Thellungiella halophila – flower/leaf transcriptomic comparison in Arabidopsis and Thellungiella. adt09-01_thellungiella – thelungiella 2 dye-swap – CATMA arrays.
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Gruenheit, N., O. Deusch, et al. (2012) Cutoffs and k-mers: implications from a transcriptome study in allopolyploid plants Bmc Genomics 13: 92
10.1186/1471-2164-13-92
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Background: Transcriptome analysis is increasingly being used to study the evolutionary origins and ecology of non-model plants. One issue for both transcriptome assembly and differential gene expression analyses is the common occurrence in plants of hybridisation and whole genome duplication (WGD) and hybridization resulting in allopolyploidy. The divergence of duplicated genes following WGD creates near identical homeologues that can be problematic for de novo assembly and also reference based assembly protocols that use short reads (35 – 100 bp). Results: Here we report a successful strategy for the assembly of two transcriptomes made using 75 bp Illumina reads from Pachycladon fastigiatum and Pachycladon cheesemanii. Both are allopolyploid plant species (2n = 20) that originated in the New Zealand Alps about 0.8 million years ago. In a systematic analysis of 19 different coverage cutoffs and 20 different k-mer sizes we showed that i) none of the genes could be assembled across all of the parameter space ii) assembly of each gene required an optimal set of parameter values and iii) these parameter values could be explained in part by different gene expression levels and different degrees of similarity between genes. Conclusions: To obtain optimal transcriptome assemblies for allopolyploid plants, k-mer size and k-mer coverage need to be considered simultaneously across a broad parameter space. This is important for assembling a maximum number of full length ESTs and for avoiding chimeric assemblies of homeologous and paralogous gene copies.
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Barker, M. S., H. Vogel, et al. (2009) Paleopolyploidy in the Brassicales: Analyses of the Cleome Transcriptome Elucidate the History of Genome Duplications in Arabidopsis and Other Brassicales Genome Biology and Evolution 1: 391-399. 10.1093/gbe/evp040
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The analysis of the Arabidopsis genome revealed evidence of three ancient polyploidy events in the evolution of the Brassicaceae, but the exact phylogenetic placement of these events is still not resolved. The most recent event is called the At-alpha (alpha) or 3R, the intermediate event is referred to as the At-beta (beta) or 2R, and the oldest is the At-gamma (gamma) or 1R. It has recently been established that At-gamma is shared with other Rosids, including papaya (Carica), poplar (Populus), and grape (Vitis), whereas data to date suggest that At-alpha is Brassicaceae specific. To address more precisely when the At-alpha and At-beta events occurred and which plant lineages share these paleopolyploidizations, we sequenced and analyzed over 4,700 normalized expressed sequence tag sequences from the Cleomaceae, the sister family to the Brassicaceae. Analysis of these Cleome data with homologous sequences from other Rosid genomes (Arabidopsis, Carica, Gossypium, Populus, and Vitis) yielded three major findings: 1) confirmation of a Cleome-specific paleopolyploidization (Cs-alpha) that is independent of the Brassicaceae At-alpha paleopolyploidization; 2) Cleome and Arabidopsis share the At-beta duplication, which is lacking from papaya within the Brassicales; and 3) rates of molecular evolution are faster for the herbaceous annual taxa Arabidopsis and Cleome than the other predominantly woody perennial Rosid lineages. These findings contribute to our understanding of the dynamics of genome duplication and evolution within one of the most comprehensively surveyed clades of plants, the Rosids, and clarify the complex history of the At-alpha, At-beta, and At-gamma duplications of Arabidopsis.
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Zhang, Y. Y., J. B. Lai, et al. (2008) Comparison analysis of transcripts from the halophyte Thellungiella halophila Journal of Integrative Plant Biology 50: 1327-1335. 10.1111/j.1744-7909.2008.00740.x
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The Brassicaceae family halophyte Thellungiella halophila has a high salinity tolerance and serves as a valuable halophytic genetic model plant with experimental convenience similar to Arabidopsis thaliana. A cDNA library of Thellungiella was generated from salt-treated seedlings including rosettes and roots. More than 1 000 randomly selected clones were sequenced and 946 expressed sequence tags (ESTs) were generated. The accession numbers of our EST data are available online in the GenBank database from EC598928 to EC599965. In total 679 unique clusters were assembled, and 632 (93%) had BLASTX hits in the nr databases and 7% are Thellungiella unique. According to the Gene Ontology (GO) hierarchy, 385 of 679 unigenes were categorized. Compared with public Arabidopsis microarray data, our results provide more potential salt tolerance genes in Thellungiella. These results will provide a broader coverage into Thellungiella transcriptome and benefit the discovery of salt tolerance related genes.
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Broadley, M. R., P. J. White, et al. (2008) Evidence of neutral transcriptome evolution in plants New Phytologist 180: 587-593. 10.1111/j.1469-8137.2008.02640.x
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The transcriptome of an organism is its set of gene transcripts (mRNAs) at a defined spatial and temporal locus. Because gene expression is affected markedly by environmental and developmental perturbations, it is widely assumed that transcriptome divergence among taxa represents adaptive phenotypic selection. This assumption has been challenged by neutral theories which propose that stochastic processes drive transcriptome evolution. To test for evidence of neutral transcriptome evolution in plants, we quantified 18 494 gene transcripts in nonsenescent leaves of 14 taxa of Brassicaceae using robust cross-species transcriptomics which includes a two-step physical and in silico-based normalization procedure based on DNA similarity among taxa. Transcriptome divergence correlates positively with evolutionary distance between taxa and with variation in gene expression among samples. Results are similar for pseudogenes and chloroplast genes evolving at different rates. Remarkably, variation in transcript abundance among root-cell samples correlates positively with transcriptome divergence among root tissues and among taxa. Because neutral processes affect transcriptome evolution in plants, many differences in gene expression among or within taxa may be nonfunctional, reflecting ancestral plasticity and founder effects. Appropriate null models are required when comparing transcriptomes in space and time.
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Hammond, J. P., H. C. Bowen, et al. (2006) A comparison of the Thlaspi caerulescens and Thlaspi arvense shoot transcriptomes New Phytologist 170: 239-260. 10.1111/j.1469-8137.2006.01662.x
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Whole-genome transcriptome profiling is revealing how biological systems are regulated at the transcriptional level. This study reports the development of a robust method to profile and compare the transcriptomes of two nonmodel plant species, Thlaspi caerulescens, a zinc (Zn) hyperaccumulator, and Thlaspi arvense, a nonhyperaccumulator, using Affymetrix Arabidopsis thaliana ATH1-121501 GeneChip (R) arrays (Affymetrix, Santa Clara, CA, USA). Transcript abundance was quantified in the shoots of agar- and compost-grown plants of both species. Analyses were optimized using a genomic DNA (gDNA)-based probe-selection strategy based on the hybridization efficiency of Thlaspi gDNA with corresponding A. thaliana probes. In silico alignments of GeneChip (R) probes with Thlaspi gene sequences, and quantitative real-time PCR, confirmed the validity of this approach. Approximately 5000 genes were differentially expressed in the shoots of T. caerulescens compared with T. arvense, including genes involved in Zn transport and compartmentalization. Future functional analyses of genes identified as differentially expressed in the shoots of these closely related species will improve our understanding of the molecular mechanisms of Zn hyperaccumulation.
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Pires, J. C., J. W. Zhao, et al. (2004) Flowering time divergence and genomic rearrangements in resynthesized Brassica polyploids (Brassicaceae) Biological Journal of the Linnean Society 82: 675-688. 10.1111/j.1095-8312.2004.00350.x
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Novel phenotypes often arise in generations immediately following polyploidization. Previous studies have shown that separate lineages derived from a resynthesized Brassica napus allopolyploid rapidly evolved heritable differences in flowering time. These early-flowering and late-flowering polyploid lines were expected to be genetically identical because they were derived from a single, chromosome-doubled amphihaploid plant. In this study, we investigated the molecular genetic basis for these flowering time differences. We assessed the diploid B. rapa and B. oleracea parents and the early- and late-flowering B. napus lineages for changes in genome structure, and for changes in transcript levels of four sets of FLOWERING LOCUS C (FLC) genes. No such changes were observed for BnFLC1, but we detected chromosomal rearrangements (e.g. de novo non-reciprocal transpositions) and changes in transcript level for BnFLC2 and BnFLC3 between the early- and late-flowering B. nap us. A chromosomal rearrangement of a genomic segment containing BnFLC3 was responsible for 29% of the phenotypic variation among the B. napus lines. Expression of BnFLC5 was silenced in all polyploids, although no changes in genome structure were detected. An ongoing investigation of 50 identical B. napus allopolyploids may further reveal the dynamics of changes in phenotype, genome and transcriptome at the early stages in polyploid evolution. (C) 2004 The Linnean Society of London.
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