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Deep sequencing uncovers commonality in small RNA profiles between transgene-induced and naturally occurring RNA silencing of chalcone synthase-A gene in petunia

Introduction of a transgene that transcribes RNA homologous to an endogenous gene in the plant genome can induce silencing of both genes, a phenomenon termed cosuppression.
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Abstract (provisional)Background

Introduction of a transgene that transcribes RNA homologous to an endogenous gene in the plant genome can induce silencing of both genes, a phenomenon termed cosuppression. Cosuppression was first discovered in transgenic petunia plants transformed with the CHS-A gene encoding chalcone synthase, in which nonpigmented sectors in flowers or completely white flowers are produced. Some of the flower-color patterns observed in transgenic petunias having CHS-A cosuppression resemble those in existing nontransgenic varieties. Although the mechanism by which white sectors are generated in nontransgenic petunia is known to be due to RNA silencing of the CHS-A gene as in cosuppression, whether the same trigger(s) and/or pattern of RNA degradation are involved in these phenomena has not been known. Here, we addressed this question using deep-sequencing and bioinformatic analyses of small RNAs.

Results

We analyzed short interfering RNAs (siRNAs) produced in nonpigmented sectors of petal tissues in transgenic petunia plants that have CHS-A cosuppression and a nontransgenic petunia variety Red Star, that has naturally occurring CHS-A RNA silencing. In both silencing systems, 21-nt and 22-nt siRNAs were the most and the second-most abundant size classes, respectively. CHS-A siRNA production was confined to exon 2, indicating that RNA degradation through the RNA silencing pathway occurred in this exon. Common siRNAs were detected in cosuppression and naturally occurring RNA silencing, and their ranks based on the number of siRNAs in these plants were correlated with each other. Noticeably, highly abundant siRNAs were common in these systems. Phased siRNAs were detected in multiple phases at multiple sites, and some of the ends of the regions that produced phased siRNAs were conserved.

Conclusions

The features of siRNA production found to be common to cosuppression and naturally occurring silencing of the CHS-A gene indicate mechanistic similarities between these silencing systems especially in the biosynthetic processes of siRNAs including cleavage of CHS-A transcripts and subsequent production of secondary siRNAs in exon 2. The data also suggest that these events occurred at multiple sites, which can be a feature of these silencing phenomena.

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Transcriptome variation along bud development in grapevine (Vitis vinifera L.)

Transcriptome variation along bud development in grapevine (Vitis vinifera L.) | Plant Genomics | Scoop.it

Abstract (provisional)

Background

Vegetative buds provide plants in temperate environments the possibility for growth and reproduction when environmental conditions are favorable. In grapevine, crucial developmental events take place within buds during two growing seasons in consecutive years. The first season, the shoot apical meristem within the bud differentiates all the basic elements of the shoot including flowering transition in lateral primordia and development of inflorescence primordia. These events practically end with bud dormancy. The second season, buds resume shoot growth associated to flower formation and development. Gene expression has been previously monitored at specific stages of bud development but has never been followed along the two growing seasons.

Results

Gene expression changes were analyzed along the bud annual cycle at eight different time points. Principal Components Analysis (PCA) revealed that the main factors explaining the global gene expression differences were the processes of bud dormancy and active growth as well as stress responses. Accordingly, non dormant buds showed an enrichment in functional categories typical of actively proliferating and growing cells together with the over abundance of transcripts belonging to stress response pathways. Differential expression analyses performed between consecutive time points indicated that major transcriptional changes were associated to para/endodormancy, endo/ecodormancy and ecodormancy/bud break transitions. Transcripts encoding key regulators of reproductive development were grouped in three major expression clusters corresponding to: (i) transcripts associated to flowering induction, (ii) transcripts associated to flower meristem specification and initiation and (iii) transcripts putatively involved in dormancy. Within this cluster, a MADS-box gene (VvFLC2) and other transcripts with similar expression patterns could participate in dormancy regulation.

Conclusions

This work provides a global view of major transcriptional changes taking place along bud development in grapevine, highlighting those molecular and biological functions involved in the main events of bud development. As reported in other woody species, the results suggest that genes regulating flowering could also be involved in dormancy regulatory pathways in grapevine.

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Construction of a high-density genetic map using specific length amplified fragment markers and identification of a quantitative trait locus for anthracnose resistance in...

Walnut (Juglans regia, 2n = 32, approximately 606 Mb per 1C genome) is an economically important tree crop. Resistance to anthracnose, caused by Colletotrichum gloeosporioides, is a major objective of walnut genetic improvement in China. The recently developed specific length amplified fragment sequencing (SLAF-seq) is an efficient strategy that can obtain large numbers of markers with sufficient sequence information to construct high-density genetic maps and permits detection of quantitative trait loci (QTLs) for molecular breeding.
Biswapriya Biswavas Misra's insight:
AbstractBackground

Walnut (Juglans regia, 2n = 32, approximately 606 Mb per 1C genome) is an economically important tree crop. Resistance to anthracnose, caused by Colletotrichum gloeosporioides, is a major objective of walnut genetic improvement in China. The recently developed specific length amplified fragment sequencing (SLAF-seq) is an efficient strategy that can obtain large numbers of markers with sufficient sequence information to construct high-density genetic maps and permits detection of quantitative trait loci (QTLs) for molecular breeding.

Results

SLAF-seq generated 161.64 M paired-end reads. 153,820 SLAF markers were obtained, of which 49,174 were polymorphic. 13,635 polymorphic markers were sorted into five segregation types and 2,577 markers of them were used to construct genetic linkage maps: 2,395 of these fell into 16 linkage groups (LGs) for the female map, 448 markers for the male map, and 2,577 markers for the integrated map. Taking into account the size of all LGs, the marker coverage was 2,664.36 cM for the female map, 1,305.58 cM for the male map, and 2,457.82 cM for the integrated map. The average intervals between two adjacent mapped markers were 1.11 cM, 2.91 cM and 0.95 cM for three maps, respectively. ‘SNP_only’ markers accounted for 89.25 % of the markers on the integrated map. Mapping markers contained 5,043 single nucleotide polymorphisms (SNPs) loci, which corresponded to two SNP loci per SLAF marker. According to the integrated map, we used interval mapping (Logarithm of odds, LOD > 3.0) to detect our quantitative trait. One QTL was detected for anthracnose resistance. The interval of this QTL ranged from 165.51 cM to 176.33 cM on LG14, and ten markers in this interval that were above the threshold value were considered to be linked markers to the anthracnose resistance trait. The phenotypic variance explained by each marker ranged from 16.2 to 19.9 %, and their LOD scores varied from 3.22 to 4.04.

Conclusions

High-density genetic maps for walnut containing 16 LGs were constructed using the SLAF-seq method with an F1 population. One QTL for walnut anthracnose resistance was identified based on the map. The results will aid molecular marker-assisted breeding and walnut resistance genes identification.

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Transcriptome analysis of the Holly mangrove Acanthus ilicifolius and its terrestrial relative, Acanthus leucostachyus, provides insights into adap...

Transcriptome analysis of the Holly mangrove Acanthus ilicifolius and its terrestrial relative, Acanthus leucostachyus, provides insights into adap... | Plant Genomics | Scoop.it
BMC Genomics. 2015 Aug 14;16(1):605. doi: 10.1186/s12864-015-1813-9.
Biswapriya Biswavas Misra's insight:
AbstractBACKGROUND:

Acanthus is a unique genus consisting of both true mangrove and terrestrial species; thus, it represents an ideal system for studying the origin and adaptive evolution of mangrove plants to intertidal environments. However, little is known regarding the two respects of mangrove species in Acanthus. In this study, we sequenced the transcriptomes of the pooled roots and leaves tissues for a mangrove species, Acanthus ilicifolius, and its terrestrial congener, A. leucostachyus, to illustrate the origin of the mangrove species in this genus and their adaptive evolution to harsh habitats.

RESULTS:

We obtained 73,039 and 69,580 contigs with N50 values of 741 and 1557 bp for A. ilicifolius and A. leucostachyus, respectively. Phylogenetic analyses based on four nuclear segments and three chloroplast fragments revealed that mangroves and terrestrial species in Acanthus fell into different clades, indicating a single origin of the mangrove species in Acanthus. Based on 6634 orthologs, A. ilicifolius and A. leucostachyus were found to be highly divergent, with a peak of synonymous substitution rate (Ks) distribution of 0.145 and an estimated divergence time of approximately 16.8 million years ago (MYA). The transgression in the Early to Middle Miocene may be the major reason for the entry of the mangrove lineage of Acanthus into intertidal environments. Gene ontology (GO) classifications of the full transcriptomes did not show any apparent differences between A. ilicifolius and A. leucostachyus, suggesting the absence of gene components specific to the mangrove transcriptomes. A total of 99 genes in A. ilicifolius were identified with signals of positive selection. Twenty-three of the 99 positively selected genes (PSGs) were found to be involved in salt, heat and ultraviolet stress tolerance, seed germination and embryo development under periodic inundation. These stress-tolerance related PSGs may be crucial for the adaptation of the mangrove species in this genus to stressful marine environments and may contribute to speciation in Acanthus.

CONCLUSIONS:

We characterized the transcriptomes of one mangrove species of Acanthus, A. ilicifolius, and its terrestrial relative, A. leucostachyus, and provided insights into the origin of the mangrove Acanthus species and their adaptive evolution to abiotic stresses in intertidal environments.

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Changes in the common bean (Phaseolus vulgaris) transcriptome in response to secreted and surface signal molecules of Rhizobium etli.

Changes in the common bean (Phaseolus vulgaris) transcriptome in response to secreted and surface signal molecules of Rhizobium etli. | Plant Genomics | Scoop.it
Establishment of nitrogen fixing symbiosis requires the recognition of rhizobial molecules to initiate the development of nodules. Using transcriptional profiling of roots inoculated with mutant strains defective in the synthesis of Nod Factor (NF), exopolysaccharide (EPS) or lipopolysaccharide (LPS), we identified 2606 genes from common bean (Phaseolus vulgaris) that are differentially regulated at early stages of its interaction with Rhizobium etli. Many transcription factors from different families are modulated by NF, EPS and LPS in different combinations, suggesting that the plant response depends on the integration of multiple signals. Some receptors identified as differentially expressed constitute excellent candidates to participate in signal perception of molecules derived from the bacteria. Several components of the ethylene signal response, a hormone that plays a negative role during early stages of the process, were down-regulated by NF and LPS. In addition, genes encoding proteins involved in small RNA-mediated gene regulation were regulated by these signal molecules, such as Argonaute 7, a specific component of the TAS3 derived tasiRNAs biosynthetic pathway, an RNA dependent RNA polymerase and a XH/XP domain-containing protein, which is part of the RNA directed-DNA methylation. Interestingly, a number of genes encoding components of the circadian central oscillator were down-regulated by NF and LPS, suggesting that a root circadian clock is adjusted at early stages of symbiosis. Our results reveal a complex interaction of the responses triggered by NF, LPS and EPS that integrates information of the signals present in the surface or secreted by rhizobia.
Biswapriya Biswavas Misra's insight:

Establishment of nitrogen fixing symbiosis requires the recognition of rhizobial molecules to initiate the development of nodules. Using transcriptional profiling of roots inoculated with mutant strains defective in the synthesis of Nod Factor (NF), exopolysaccharide (EPS) or lipopolysaccharide (LPS), we identified 2606 genes from common bean (Phaseolus vulgaris) that are differentially regulated at early stages of its interaction with Rhizobium etli. Many transcription factors from different families are modulated by NF, EPS and LPS in different combinations, suggesting that the plant response depends on the integration of multiple signals. Some receptors identified as differentially expressed constitute excellent candidates to participate in signal perception of molecules derived from the bacteria. Several components of the ethylene signal response, a hormone that plays a negative role during early stages of the process, were down-regulated by NF and LPS. In addition, genes encoding proteins involved in small RNA-mediated gene regulation were regulated by these signal molecules, such as Argonaute 7, a specific component of the TAS3 derived tasiRNAs biosynthetic pathway, an RNA dependent RNA polymerase and a XH/XP domain-containing protein, which is part of the RNA directed-DNA methylation. Interestingly, a number of genes encoding components of the circadian central oscillator were down-regulated by NF and LPS, suggesting that a root circadian clock is adjusted at early stages of symbiosis. Our results reveal a complex interaction of the responses triggered by NF, LPS and EPS that integrates information of the signals present in the surface or secreted by rhizobia.

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Genome-wide annotation and characterization of CLAVATA/ESR (CLE) peptide hormones of soybean (Glycine max) and common bean (Phaseolus vulgaris), and their orthologues of Arabidopsis thaliana

CLE peptides are key regulators of cell proliferation and differentiation in plant shoots, roots, vasculature, and legume nodules. They are C-terminally encoded peptides that are post-translationally cleaved and modified from their corresponding pre-propeptides to produce a final ligand that is 12–13 amino acids in length. In this study, an array of bionformatic and comparative genomic approaches was used to identify and characterize the complete family of CLE peptide-encoding genes in two of the world’s most important crop species, soybean and common bean. In total, there are 84 CLE peptide-encoding genes in soybean (considerably more than the 32 present in Arabidopsis), including three pseudogenes and two multi-CLE domain genes having six putative CLE domains each. In addition, 44 CLE peptide-encoding genes were identified in common bean. In silico characterization was used to establish all soybean homeologous pairs, and to identify corresponding gene orthologues present in common bean and Arabidopsis. The soybean CLE pre-propeptide family was further analysed and separated into seven distinct groups based on structure, with groupings strongly associated with the CLE domain sequence and function. These groups provide evolutionary insight into the CLE peptide families of soybean, common bean, and Arabidopsis, and represent a novel tool that can aid in the functional characterization of the peptides. Transcriptional evidence was also used to provide further insight into the location and function of all CLE peptide-encoding members currently available in gene atlases for the three species. Taken together, this in-depth analysis helped to identify and categorize the complete CLE peptide families of soybean and common bean, established gene orthologues within the two legume species, and Arabidopsis, and provided a platform to help compare, contrast, and identify the function of critical CLE peptide hormones in plant development.

Via Christophe Jacquet
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Transcriptomic profiling of sequential tumours from breast cancer patients provides a global view of metastatic expression changes following endocrine therapy

Transcriptomic profiling of sequential tumours from breast cancer patients provides a global view of metastatic exp http://t.co/DZQTjiTZTs
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Transcriptomic Characterization of Soybean Roots in Response to Bradyrhizobium Infection by RNA Sequencing

Legumes interact with rhizobium convert N2 into ammonia for plant use. To investigate the plant basal nitrogen fixation mechanisms induced in response to Bradyrhizobium, differential gene expression in root of inoculated and mock-inoculated soybean was analysed by RNA-Seq. A total of 55787 transcripts were aligned to soybean genome reference sequences, 280 and 316 transcripts were found to be up- and down-regulated, respectively, in inoculated relative to mock-inoculated soybean???s root at V1 stage. Gene ontology (GO) analyses detected 104, 182 and 178 genes associated with cell component category, molecular function category and biological process category, respectively. Pathway analysis revealed that 98 differentiallly expressed genes (115 transcripts) involved in 169 biological pathways. We selected 19 differentially expressed genes and analyzed their expressions in mock-inoculated, inoculated USDA110 and CCBAU45436 using qRT-PCR. The results were consistent with those obtained from rhizobia infected RNA-Seq data. These showed that the results of RNA-Seq have reliability and universality. Additionally, this study showed some novel genes associated with nitrogen fixation process comparison with the previously identified QTLs.
Biswapriya Biswavas Misra's insight:

Legumes interact with rhizobium convert N2 into ammonia for plant use. To investigate the plant basal nitrogen fixation mechanisms induced in response to Bradyrhizobium, differential gene expression in root of inoculated and mock-inoculated soybean was analysed by RNA-Seq. A total of 55787 transcripts were aligned to soybean genome reference sequences, 280 and 316 transcripts were found to be up- and down-regulated, respectively, in inoculated relative to mock-inoculated soybean???s root at V1 stage. Gene ontology (GO) analyses detected 104, 182 and 178 genes associated with cell component category, molecular function category and biological process category, respectively. Pathway analysis revealed that 98 differentiallly expressed genes (115 transcripts) involved in 169 biological pathways. We selected 19 differentially expressed genes and analyzed their expressions in mock-inoculated, inoculated USDA110 and CCBAU45436 using qRT-PCR. The results were consistent with those obtained from rhizobia infected RNA-Seq data. These showed that the results of RNA-Seq have reliability and universality. Additionally, this study showed some novel genes associated with nitrogen fixation process comparison with the previously identified QTLs.

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Transformation of Ulva mutabilis (Chlorophyta) by vector plasmids integrating into the genome

Transformation of Ulva mutabilis (Chlorophyta) by vector plasmids integrating into the genome | Plant Genomics | Scoop.it

A method for the stable transformation of the green marine macroalga Ulva mutabilis was developed based on vector plasmids integrating into the genome. By combination of the expression signals (promoter, enhancer and transcriptional termination sequences) of a chromosomal rbcS gene from Ulva mutabilis with the bleomycin resistance gene (ble) from Streptoalloteichus hindustanus, a dominant selectable marker gene was constructed for the preparation of a series of E. coli - Ulva mutabilis shuttle vector plasmids. Special vectors were prepared for the introduction and expression of foreign genes in Ulva, for insertional mutagenesis and gene tagging by plasmid integration into the genome, and for protein-tagging by the green fluorescent protein (GFP), as well as tools for post-transcriptional gene silencing and cosmid cloning to prepare genomic gene libraries for mutant gene complementation. The vectors were successfully tested in pilot experiments, where they were efficiently introduced into Ulva gametes, zoospores or protoplasts of somatic blade cells by treatment with Ca++-ions and polyethylene glycol under isotonic conditions at low ionic strength. The parthenogenetically propagated phleomycin resistant transformants of the mutant slender (sl) and the wildtype (wt) were demonstrated to be carrying the plasmids randomly, which were stably integrated into the chromosomes, often as tandem repeat clusters.


Via Pierre-Marc Delaux
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A CRISPR/Cas9 toolbox for multiplexed plant genome editing and transcriptional regulation

A CRISPR/Cas9 toolbox for multiplexed plant genome editing and transcriptional regulation | Plant Genomics | Scoop.it
Biswapriya Biswavas Misra's insight:

The relative ease, speed and biological scope of CRISPR/Cas9-based reagents for genomic manipulations are revolutionizing virtually all areas of molecular biosciences, including functional genomics, genetics, applied biomedical research and agricultural biotechnology. In plant systems, however, a number of hurdles currently exist that limit this technology from reaching its full potential. For example, significant plant molecular biology expertise and effort is still required to generate functional expression constructs that allow simultaneous editing, and especially transcriptional regulation, of multiple different genomic loci or "multiplexing", which is a significant advantage of CRISPR/Cas9 versus other genome editing systems. In order to streamline and facilitate rapid and wide-scale use of CRISPR/Cas9-based technologies for plant research, we developed and implemented a comprehensive molecular toolbox for multifaceted CRISPR/Cas9 applications in plants. This toolbox provides researchers with a protocol and reagents to quickly and efficiently assemble functional CRISPR/Cas9 T-DNA constructs for monocots and dicots using Golden Gate and Gateway cloning methods. It comes with a full suite of capabilities, including multiplexed gene editing and transcriptional activation or repression of plant endogenous genes. We report the functionality and effectiveness of this toolbox in model plants such as tobacco, Arabidopsis and rice, demonstrating its utility for basic and applied plant research.

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The environment exerts a greater influence than the transgene on the transcriptome of field-grown wheat expressing the Pm3b allele.

The environment exerts a greater influence than the transgene on the transcriptome of field-grown wheat expressing the Pm3b allele. | Plant Genomics | Scoop.it
Wheat provides 20 % of the calories consumed worldwide. Powdery mildew infections of wheat can result in more than 30 % yield loss but it has been demonstrated that wheat overexpressing Pm3b, an allele of the R gene Pm3, has enhanced resistance against powdery mildew under field conditions. A gene expression profile study using GeneChip Wheat Genome Array and Fluidigm 96.96 Dynamic Arrays was performed to obtain insights into the mode of action of Pm3b and to elucidate the molecular basis of pleiotropic effects observed in three out of four independent transgenic events under field conditions. A cluster analysis of the microarray data and a principal component analysis of the Fluidigm 96.96 Dynamic Arrays data showed that transgenic lines and null segregants grouped together. The microarray analysis of samples from fungicide-treated plants revealed that significantly fewer genes were differentially expressed in Pm3b#1 than in Pm3b#2, which had a pleiotropic phenotype in the field, compared to their null segregants. Together, our data provide evidence that the environment influenced gene expression in the Pm3b lines more than the transgene itself.
Biswapriya Biswavas Misra's insight:

Wheat provides 20 % of the calories consumed worldwide. Powdery mildew infections of wheat can result in more than 30 % yield loss but it has been demonstrated that wheat overexpressing Pm3b, an allele of the R gene Pm3, has enhanced resistance against powdery mildew under field conditions. A gene expression profile study using GeneChip Wheat Genome Array and Fluidigm 96.96 Dynamic Arrays was performed to obtain insights into the mode of action of Pm3b and to elucidate the molecular basis of pleiotropic effects observed in three out of four independent transgenic events under field conditions. A cluster analysis of the microarray data and a principal component analysis of the Fluidigm 96.96 Dynamic Arrays data showed that transgenic lines and null segregants grouped together. The microarray analysis of samples from fungicide-treated plants revealed that significantly fewer genes were differentially expressed in Pm3b#1 than in Pm3b#2, which had a pleiotropic phenotype in the field, compared to their null segregants. Together, our data provide evidence that the environment influenced gene expression in the Pm3b lines more than the transgene itself.

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Development of genome-wide insertion and deletion markers for maize, based on next-generation sequencing data

Background

Insertions and deletions (indels) are the most abundant form of structural variation in all genomes. Indels have been increasingly recognized as an important source of molecular markers due to high-density occurrence, cost-effectiveness, and ease of genotyping. Coupled with developments in bioinformatics, next-generation sequencing (NGS) platforms enable the discovery of millions of indel polymorphisms by comparing the whole genome sequences of individuals within a species.
Results

A total of 1,973,746 unique indels were identified in 345 maize genomes, with an overall density of 958.79 indels/Mbp, and an average allele number of 2.76, ranging from 2 to 107. There were 264,214 indels with polymorphism information content (PIC) values greater than or equal to 0.5, accounting for 13.39 % of overall indels. Of these highly polymorphic indels, we designed primer pairs for 83,481 and 29,403 indels with major allele differences (i.e. the size difference between the most and second most frequent alleles) greater than or equal to 3 and 8 bp, respectively, based on the differing resolution capabilities of gel electrophoresis. The accuracy of our indel markers was experimentally validated, and among 100 indel markers, average accuracy was approximately 90 %. In addition, we also validated the polymorphism of the indel markers. Of 100 highly polymorphic indel markers, all had polymorphisms with average PIC values of 0.54.
Conclusions

The maize genome is rich in indel polymorphisms. Intriguingly, the level of polymorphism in genic regions of the maize genome was higher than that in intergenic regions. The polymorphic indel markers developed from this study may enhance the efficiency of genetic research and marker-assisted breeding in maize.
Biswapriya Biswavas Misra's insight:
Background

Insertions and deletions (indels) are the most abundant form of structural variation in all genomes. Indels have been increasingly recognized as an important source of molecular markers due to high-density occurrence, cost-effectiveness, and ease of genotyping. Coupled with developments in bioinformatics, next-generation sequencing (NGS) platforms enable the discovery of millions of indel polymorphisms by comparing the whole genome sequences of individuals within a species.

Results

A total of 1,973,746 unique indels were identified in 345 maize genomes, with an overall density of 958.79 indels/Mbp, and an average allele number of 2.76, ranging from 2 to 107. There were 264,214 indels with polymorphism information content (PIC) values greater than or equal to 0.5, accounting for 13.39 % of overall indels. Of these highly polymorphic indels, we designed primer pairs for 83,481 and 29,403 indels with major allele differences (i.e. the size difference between the most and second most frequent alleles) greater than or equal to 3 and 8 bp, respectively, based on the differing resolution capabilities of gel electrophoresis. The accuracy of our indel markers was experimentally validated, and among 100 indel markers, average accuracy was approximately 90 %. In addition, we also validated the polymorphism of the indel markers. Of 100 highly polymorphic indel markers, all had polymorphisms with average PIC values of 0.54.

Conclusions

The maize genome is rich in indel polymorphisms. Intriguingly, the level of polymorphism in genic regions of the maize genome was higher than that in intergenic regions. The polymorphic indel markers developed from this study may enhance the efficiency of genetic research and marker-assisted breeding in maize.

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Uncovering co-expression gene network modules regulating fruit acidity in diverse apples

Acidity is a major contributor to fruit quality. Several organic acids are present in apple fruit, but malic acid is predominant and determines fruit acidity. The trait is largely controlled by the Malic acid (Ma) locus, underpinning which Ma1 that putatively encodes a vacuolar aluminum-activated malate transporter1 (ALMT1)-like protein is a strong candidate gene. We hypothesize that fruit acidity is governed by a gene network in which Ma1 is key member. The goal of this study is to identify the gene network and the potential mechanisms through which the network operates.
Biswapriya Biswavas Misra's insight:
Background

Acidity is a major contributor to fruit quality. Several organic acids are present in apple fruit, but malic acid is predominant and determines fruit acidity. The trait is largely controlled by the Malic acid (Ma) locus, underpinning which Ma1 that putatively encodes a vacuolar aluminum-activated malate transporter1 (ALMT1)-like protein is a strong candidate gene. We hypothesize that fruit acidity is governed by a gene network in which Ma1 is key member. The goal of this study is to identify the gene network and the potential mechanisms through which the network operates.

Results

Guided by Ma1, we analyzed the transcriptomes of mature fruit of contrasting acidity from six apple accessions of genotype Ma_ (MaMa or Mama) and four of mama using RNA-seq and identified 1301 fruit acidity associated genes, among which 18 were most significant acidity genes (MSAGs). Network inferring using weighted gene co-expression network analysis (WGCNA) revealed five co-expression gene network modules of significant (P < 0.001) correlation with malate. Of these, the Ma1 containing module (Turquoise) of 336 genes showed the highest correlation (0.79). We also identified 12 intramodular hub genes from each of the five modules and 18 enriched gene ontology (GO) terms and MapMan sub-bines, including two GO terms (GO:0015979 and GO:0009765) and two MapMap sub-bins (1.3.4 and 1.1.1.1) related to photosynthesis in module Turquoise. Using Lemon-Tree algorithms, we identified 12 regulator genes of probabilistic scores 35.5–81.0, including MDP0000525602 (a LLR receptor kinase), MDP0000319170 (an IQD2-like CaM binding protein) and MDP0000190273 (an EIN3-like transcription factor) of greater interest for being one of the 18 MSAGs or one of the 12 intramodular hub genes in Turquoise, and/or a regulator to the cluster containing Ma1.

Conclusions

The most relevant finding of this study is the identification of the MSAGs, intramodular hub genes, enriched photosynthesis related processes, and regulator genes in a WGCNA module Turquoise that not only encompasses Ma1 but also shows the highest modular correlation with acidity. Overall, this study provides important insight into the Ma1-mediated gene network controlling acidity in mature apple fruit of diverse genetic background.

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Homologues of xenobiotic metabolizing N-acetyltransferases in plant-associated fungi: Novel functions for an old enzyme family

Homologues of xenobiotic metabolizing N-acetyltransferases in plant-associated fungi: Novel functions for an old enzyme family | Plant Genomics | Scoop.it
Biswapriya Biswavas Misra's insight:

Plant-pathogenic fungi and their hosts engage in chemical warfare, attacking each other with toxic products of secondary metabolism and defending themselves via an arsenal of xenobiotic metabolizing enzymes. One such enzyme is homologous to arylamine N-acetyltransferase (NAT) and has been identified in Fusarium infecting cereal plants as responsible for detoxification of host defence compound 2-benzoxazolinone. Here we investigate functional diversification of NAT enzymes in crop-compromising species of Fusarium and Aspergillus, identifying three groups of homologues: Isoenzymes of the first group are found in all species and catalyse reactions with acetyl-CoA or propionyl-CoA. The second group is restricted to the plant pathogens and is active with malonyl-CoA in Fusarium species infecting cereals. The third group generates minimal activity with acyl-CoA compounds that bind non-selectively to the proteins. We propose that fungal NAT isoenzymes may have evolved to perform diverse functions, potentially relevant to pathogen fitness, acetyl-CoA/propionyl-CoA intracellular balance and secondary metabolism.

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Intraspecific comparative analyses of metabolites between diploid and tetraploid Arabidopsis thaliana and Pyrus communis

Abstract
Background

It has been said that naturally occurring autopolyploid strains are more tolerant of biotic and/or abiotic stresses, due at least in part to the higher accumulation of secondary metabolites. Data supporting this hypothesis come from comparisons between naturally established autopolyploids and diploids; thus the high accumulation of metabolites in polyploid strains may be a secondarily acquired feature and not a direct effect of the autopolyploidy. But no detailed studies on this issue have been carried out.
Results

Here we carried out metabolome analyses between newly created tetraploids and the parent diploid in a model plant, Arabidopsis thaliana, and the agriculturally important pear fruit tree (Pyrus communis var. sativa). Our data showed that small numbers of metabolite species differ in amount between diploids and tetraploids in both species, but the differences were not reproducible among growth conditions and species.
Conclusions

These results strongly indicate that metabolite content is not universal nor the direct target of polyploidy-dependent changes. Instead, naturally occurring hyperaccumulation of metabolites in autopolyploids may be the result of secondary natural selection.
Biswapriya Biswavas Misra's insight:
AbstractBackground

It has been said that naturally occurring autopolyploid strains are more tolerant of biotic and/or abiotic stresses, due at least in part to the higher accumulation of secondary metabolites. Data supporting this hypothesis come from comparisons between naturally established autopolyploids and diploids; thus the high accumulation of metabolites in polyploid strains may be a secondarily acquired feature and not a direct effect of the autopolyploidy. But no detailed studies on this issue have been carried out.

Results

Here we carried out metabolome analyses between newly created tetraploids and the parent diploid in a model plant, Arabidopsis thaliana, and the agriculturally important pear fruit tree (Pyrus communis var. sativa). Our data showed that small numbers of metabolite species differ in amount between diploids and tetraploids in both species, but the differences were not reproducible among growth conditions and species.

Conclusions

These results strongly indicate that metabolite content is not universal nor the direct target of polyploidy-dependent changes. Instead, naturally occurring hyperaccumulation of metabolites in autopolyploids may be the result of secondary natural selection.

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Identification of in vitro and in vivo disconnects using transcriptomic data

Integrating transcriptomic experiments within drug development is increasingly advocated for the early detection of toxicity. This is partly to reduce costs related to drug failures in the late, and expensive phases of clinical trials. Such an approach has proven useful both in the study of toxicology and carcinogenicity. However, general lack of translation of in vitro findings to in vivo systems remains one of the bottle necks in drug development. This paper proposes a method for identifying disconnected genes between in vitro and in vivo toxicogenomic rat experiments. The analytical framework is based on the joint modeling of dose-dependent in vitro and in vivo data using a fractional polynomial framework and biclustering algorithm.
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De novo assembly and characterisation of the field pea transcriptome using RNA-Seq.

BMC Genomics. 2015 Aug 16;16(1):611. doi: 10.1186/s12864-015-1815-7.
Biswapriya Biswavas Misra's insight:
AbstractBACKGROUND:

Field pea (Pisum sativum L.) is a cool-season grain legume that is cultivated world-wide for both human consumption and stock-feed purposes. Enhancement of genetic and genomic resources for field pea will permit improved understanding of the control of traits relevant to crop productivity and quality. Advances in second-generation sequencing and associated bioinformatics analysis now provide unprecedented opportunities for the development of such resources. The objective of this study was to perform transcriptome sequencing and characterisation from two genotypes of field pea that differ in terms of seed and plant morphological characteristics.

RESULTS:

Transcriptome sequencing was performed with RNA templates from multiple tissues of the field pea genotypes Kaspa and Parafield. Tissue samples were collected at various growth stages, and a total of 23 cDNA libraries were sequenced using Illumina high-throughput sequencing platforms. A total of 407 and 352 million paired-end reads from the Kaspa and Parafield transcriptomes, respectively were assembled into 129,282 and 149,272 contigs, which were filtered on the basis of known gene annotations, presence of open reading frames (ORFs), reciprocal matches and degree of coverage. Totals of 126,335 contigs from Kaspa and 145,730 from Parafield were subsequently selected as the reference set. Reciprocal sequence analysis revealed that c. 87 % of contigs were expressed in both cultivars, while a small proportion were unique to each genotype. Reads from different libraries were aligned to the genotype-specific assemblies in order to identify and characterise expression of contigs on a tissue-specific basis, of which 87 % were expressed in more than one tissue, while others showed distinct expression patterns in specific tissues, providing unique transcriptome signatures.

CONCLUSION:

This study provided a comprehensive assembled and annotated transcriptome set for field pea that can be used for development of genetic markers, in order to assess genetic diversity, construct linkage maps, perform trait-dissection and implement whole-genome selection strategies in varietal improvement programs, as well to identify target genes for genetic modification approaches on the basis of annotation and expression analysis. In addition, the reference field pea transcriptome will prove highly valuable for comparative genomics studies and construction of a finalised genome sequence.

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Expanding the repertoire of secretory peptides controlling root development with comparative genome analysis and functional assays

Expanding the repertoire of secretory peptides controlling root development with comparative genome analysis and functional assays | Plant Genomics | Scoop.it

Small peptides of the Arabidopsis GLV/RGF/CLEL family are involved in different developmental programmes, including meristem maintenance and gravitropic responses. In addition, our previous report suggested that they also participate in the formation of lateral roots. Specifically, GLV6 is transcribed during the first stages of primordium development and GLV6 overexpression results in a strong reduction of emerged lateral roots. To investigate the cause of this phenotype we analysed primordium development in gain-of-function (gof) mutants and found that GLV6 induces supernumerary pericycle divisions, hindering the formation of a dome-shaped primordium, a prerequisite for successful emergence. The GLV6 phenotype could be reproduced by ectopic expression of the gene only in xylem-pole pericycle cells. Furthermore, GLV6 seems to function at the very beginning of lateral root initiation because GLV6 excess—either gene overexpression or peptide treatment—disrupts the first asymmetric cell divisions required for proper primordium formation. Our results suggest that GLV6 acts during lateral root initiation controlling the patterning of the first pericycle divisions.


Via Christophe Jacquet
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Cell-specific transcriptomic analyses of three-dimensional shoot development in the moss Physcomitrella patens.

Cell-specific transcriptomic analyses of three-dimensional shoot development in the moss Physcomitrella patens. | Plant Genomics | Scoop.it
Plant J. 2015 Aug;83(4):743-751. doi: 10.1111/tpj.12928. Epub 2015 Jul 22.
Biswapriya Biswavas Misra's insight:

Haploid moss gametophytes harbor distinct stem cell types, including tip cells that divide in single planes to generate filamentous protonemata, and bud cells that divide in three planes to yield axial gametophore shoots. This transition from filamentous to triplanar growth occurs progressively during the moss life cycle, and is thought to mirror evolution of the first terrestrial plants from Charophycean green algal ancestors. The innovation of morphologically complex plant body plans facilitated colonization of the vertical landscape, and enabled development of complex vegetative and reproductive plant morphologies. Despite its profound evolutionary significance, the molecular programs involved in this transition from filamentous to triplanar meristematic plant growth are poorly understood. In this study, we used single-cell type transcriptomics to identify more than 4000 differentially expressed genes that distinguish uniplanar protonematal tip cells from multiplanar gametophore bud cells in the moss Physcomitrella patens. While the transcriptomes of both tip and bud cells show molecular signatures of proliferative cells, the bud cell transcriptome exhibits a wider variety of genes with significantly increased transcript abundances. Our data suggest that combined expression of genes involved in shoot patterning and asymmetric cell division accompanies the transition from uniplanar to triplanar meristematic growth in moss.

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Oil biosynthesis in a basal angiosperm: transcriptome analysis of Persea Americana mesocarp

Oil biosynthesis in a basal angiosperm: transcriptome analysis of Persea Americana mesocarp | Plant Genomics | Scoop.it

Via Andres Zurita
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Andres Zurita's curator insight, August 19, 4:47 PM
Background

The mechanism by which plants synthesize and store high amounts of triacylglycerols (TAG) in tissues other than seeds is not well understood. The comprehension of controls for carbon partitioning and oil accumulation in nonseed tissues is essential to generate oil-rich biomass in perennial bioenergy crops. Persea americana (avocado), a basal angiosperm with unique features that are ancestral to most flowering plants, stores ~ 70 % TAG per dry weight in its mesocarp, a nonseed tissue. Transcriptome analyses of select pathways, from generation of pyruvate and leading up to TAG accumulation, in mesocarp tissues of avocado was conducted and compared with that of oil-rich monocot (oil palm) and dicot (rapeseed and castor) tissues to identify tissue- and species-specific regulation and biosynthesis of TAG in plants.

Results

RNA-Seq analyses of select lipid metabolic pathways of avocado mesocarp revealed patterns similar to that of other oil-rich species. However, only some predominant orthologs of the fatty acid biosynthetic pathway genes in this basal angiosperm were similar to those of monocots and dicots. The accumulation of TAG, rich in oleic acid, was associated with higher transcript levels for a putative stearoyl-ACP desaturase and endoplasmic reticulum (ER)-associated acyl-CoA synthetases, during fruit development. Gene expression levels for enzymes involved in terminal steps to TAG biosynthesis in the ER further indicated that both acyl-CoA-dependent and -independent mechanisms might play a role in TAG assembly, depending on the developmental stage of the fruit. Furthermore, in addition to the expression of an ortholog of WRINKLED1 (WRI1), a regulator of fatty acid biosynthesis, high transcript levels for WRI2-like and WRI3-like suggest a role for additional transcription factors in nonseed oil accumulation. Plastid pyruvate necessary for fatty acid synthesis is likely driven by the upregulation of genes involved in glycolysis and transport of its intermediates. Together, a comparative transcriptome analyses for storage oil biosynthesis in diverse plants and tissues suggested that several distinct and conserved features in this basal angiosperm species might contribute towards its rich TAG content.

Conclusions

Our work represents a comprehensive transcriptome resource for a basal angiosperm species and provides insight into their lipid metabolism in mesocarp tissues. Furthermore, comparison of the transcriptome of oil-rich mesocarp of avocado, with oil-rich seed and nonseed tissues of monocot and dicot species, revealed lipid gene orthologs that are highly conserved during evolution. The orthologs that are distinctively expressed in oil-rich mesocarp tissues of this basal angiosperm, such as WRI2, ER-associated acyl-CoA synthetases, and lipid-droplet associated proteins were also identified. This study provides a foundation for future investigations to increase oil-content and has implications for metabolic engineering to enhance storage oil content in nonseed tissues of diverse species.

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Transcriptomic Analysis of Sinorhizobium meliloti and Medicago truncatula Symbiosis Using Nitrogen Fixation–Deficient Nodules

Transcriptomic Analysis of Sinorhizobium meliloti and Medicago truncatula Symbiosis Using Nitrogen Fixation–Deficient Nodules | Plant Genomics | Scoop.it
The bacterium Sinorhizobium meliloti interacts symbiotically with legume plant hosts such as Medicago truncatula to form nitrogen-fixing root nodules. During symbiosis, plant and bacterial cells differentiate in a coordinated manner, resulting in specialized plant cells that contain nitrogen-fixing bacteroids. Both plant and bacterial genes are required at each developmental stage of symbiosis. We analyzed gene expression in nodules formed by wild-type bacteria on six plant mutants with defects in nitrogen fixation. We observed differential expression of 482 S. meliloti genes with functions in cell envelope homeostasis, cell division, stress response, energy metabolism, and nitrogen fixation. We simultaneously analyzed gene expression in M. truncatula and observed differential regulation of host processes that may trigger bacteroid differentiation and control bacterial infection. Our analyses of developmentally arrested plant mutants indicate that plants use distinct means to control bacterial infection during early and late symbiotic stages.

Via Christophe Jacquet
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Genome-wide association mapping reveals novel sources of resistance to northern corn leaf blight in maize

Genome-wide association mapping reveals novel sources of resistance to northern corn leaf blight in maize | Plant Genomics | Scoop.it
Background

Northern corn leaf blight (NCLB) caused by Exserohilum turcicum is a destructive disease in maize. Using host resistance to minimize the detrimental effects of NCLB on maize productivity is the most cost-effective and appealing disease management strategy. However, this requires the identification and use of stable resistance genes that are effective across different environments.
Results

We evaluated a diverse maize population comprised of 999 inbred lines across different environments for resistance to NCLB. To identify genomic regions associated with NCLB resistance in maize, a genome-wide association analysis was conducted using 56,110 single-nucleotide polymorphism markers. Single-marker and haplotype-based associations, as well as Anderson-Darling tests, identified alleles significantly associated with NCLB resistance. The single-marker and haplotype-based association mappings identified twelve and ten loci (genes), respectively, that were significantly associated with resistance to NCLB. Additionally, by dividing the population into three subgroups and performing Anderson-Darling tests, eighty one genes were detected, and twelve of them were related to plant defense. Identical defense genes were identified using the three analyses.
Conclusion

An association panel including 999 diverse lines was evaluated for resistance to NCLB in multiple environments, and a large number of resistant lines were identified and can be used as reliable resistance resource in maize breeding program. Genome-wide association study reveals that NCLB resistance is a complex trait which is under the control of many minor genes with relatively low effects. Pyramiding these genes in the same background is likely to result in stable resistance to NCLB.

Via Christophe Jacquet
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The Regulatory Status of Genome-edited Crops

The Regulatory Status of Genome-edited Crops | Plant Genomics | Scoop.it
Biswapriya Biswavas Misra's insight:

Genome editing with engineered nucleases (GEEN) represents a highly specific and efficient tool for crop improvement with the potential to rapidly generate useful novel phenotypes/traits. Genome editing techniques initiate specifically targeted double strand breaks facilitating DNA-repair pathways that lead to base additions or deletions by non-homologous end joining as well as targeted gene replacements or transgene insertions involving homology-directed repair mechanisms. Many of these techniques and the ancillary processes they employ generate phenotypic variation that is indistinguishable from that obtained through natural means or conventional mutagenesis; and therefore, they do not readily fit current definitions of genetically engineered or genetically modified used within most regulatory regimes. Addressing ambiguities regarding the regulatory status of genome editing techniques is critical to their application for development of economically useful crop traits. Continued regulatory focus on the process used, rather than the nature of the novel phenotype developed, results in confusion on the part of regulators, product developers, and the public alike and creates uncertainty as of the use of genome engineering tools for crop improvement.

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Proteomic analysis of flooded soybean root exposed to aluminum oxide nanoparticles

Proteomic analysis of flooded soybean root exposed to aluminum oxide nanoparticles | Plant Genomics | Scoop.it
Aluminum oxide (Al2O3) nanoparticles are used in agricultural products and cause various adverse growth effects on different plant species. To study the effects of Al2O3 nanoparticles on soybean under flooding stress, a gel-free proteomic technique was used. Morphological analysis revealed that treatment with 50 ppm Al2O3 nanoparticles under flooding stress enhanced soybean growth compared to ZnO and Ag nanoparticles. A total of 172 common proteins that significantly changed in abundance among control, flooding-stressed, and flooding-stressed soybean treated with Al2O3 nanoparticles were mainly related to energy metabolism. Under Al2O3 nanoparticles the energy metabolism was decreased compared to flooding stress. Hierarchical clustering divided identified proteins into four clusters, with proteins related to glycolysis exhibiting the greatest changes in abundance. Al2O3 nanoparticle-responsive proteins were predominantly related to protein synthesis/degradation, glycolysis, and lipid metabolism. mRNA expression analysis of Al2O3 nanoparticle- responsive proteins that displayed a 5-fold change in abundance revealed that NmrA-like negative transcriptional regulator was up-regulated, and flavodoxin-like quinone reductase was down-regulated. Moreover, cell death in root including hypocotyl was less evident in flooding-stressed with Al2O3 nanoparticles compared to flooding-treated soybean. These results suggest that Al2O3 nanoparticles might promote the growth of soybean under flooding stress by regulating energy metabolism and cell death.
Biswapriya Biswavas Misra's insight:

Aluminum oxide (Al2O3) nanoparticles are used in agricultural products and cause various adverse growth effects on different plant species. To study the effects of Al2O3 nanoparticles on soybean under flooding stress, a gel-free proteomic technique was used. Morphological analysis revealed that treatment with 50 ppm Al2O3 nanoparticles under flooding stress enhanced soybean growth compared to ZnO and Ag nanoparticles. A total of 172 common proteins that significantly changed in abundance among control, flooding-stressed, and flooding-stressed soybean treated with Al2O3 nanoparticles were mainly related to energy metabolism. Under Al2O3 nanoparticles the energy metabolism was decreased compared to flooding stress. Hierarchical clustering divided identified proteins into four clusters, with proteins related to glycolysis exhibiting the greatest changes in abundance. Al2O3 nanoparticle-responsive proteins were predominantly related to protein synthesis/degradation, glycolysis, and lipid metabolism. mRNA expression analysis of Al2O3 nanoparticle- responsive proteins that displayed a 5-fold change in abundance revealed that NmrA-like negative transcriptional regulator was up-regulated, and flavodoxin-like quinone reductase was down-regulated. Moreover, cell death in root including hypocotyl was less evident in flooding-stressed with Al2O3 nanoparticles compared to flooding-treated soybean. These results suggest that Al2O3 nanoparticles might promote the growth of soybean under flooding stress by regulating energy metabolism and cell death.

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Deep sequencing of the Mexican avocado transcriptome, an ancient angiosperm with a high content of fatty acids

Background

Avocado (Persea americana) is an economically important tropical fruit considered to be a good source of fatty acids. Despite its importance, the molecular and cellular characterization of biochemical and developmental processes in avocado is limited due to the lack of transcriptome and genomic information.
Results

The transcriptomes of seeds, roots, stems, leaves, aerial buds and flowers were determined using different sequencing platforms. Additionally, the transcriptomes of three different stages of fruit ripening (pre-climacteric, climacteric and post-climacteric) were also analyzed. The analysis of the RNAseqatlas presented here reveals strong differences in gene expression patterns between different organs, especially between root and flower, but also reveals similarities among the gene expression patterns in other organs, such as stem, leaves and aerial buds (vegetative organs) or seed and fruit (storage organs). Important regulators, functional categories, and differentially expressed genes involved in avocado fruit ripening were identified. Additionally, to demonstrate the utility of the avocado gene expression atlas, we investigated the expression patterns of genes implicated in fatty acid metabolism and fruit ripening.
Conclusions

A description of transcriptomic changes occurring during fruit ripening was obtained in Mexican avocado, contributing to a dynamic view of the expression patterns of genes involved in fatty acid biosynthesis and the fruit ripening process.
Biswapriya Biswavas Misra's insight:
Background

Avocado (Persea americana) is an economically important tropical fruit considered to be a good source of fatty acids. Despite its importance, the molecular and cellular characterization of biochemical and developmental processes in avocado is limited due to the lack of transcriptome and genomic information.

Results

The transcriptomes of seeds, roots, stems, leaves, aerial buds and flowers were determined using different sequencing platforms. Additionally, the transcriptomes of three different stages of fruit ripening (pre-climacteric, climacteric and post-climacteric) were also analyzed. The analysis of the RNAseqatlas presented here reveals strong differences in gene expression patterns between different organs, especially between root and flower, but also reveals similarities among the gene expression patterns in other organs, such as stem, leaves and aerial buds (vegetative organs) or seed and fruit (storage organs). Important regulators, functional categories, and differentially expressed genes involved in avocado fruit ripening were identified. Additionally, to demonstrate the utility of the avocado gene expression atlas, we investigated the expression patterns of genes implicated in fatty acid metabolism and fruit ripening.

Conclusions

A description of transcriptomic changes occurring during fruit ripening was obtained in Mexican avocado, contributing to a dynamic view of the expression patterns of genes involved in fatty acid biosynthesis and the fruit ripening process.

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De novo assembly and characterisation of the field pea transcriptome using RNA-Seq

Field pea (Pisum sativum L.) is a cool-season grain legume that is cultivated world-wide for both human consumption and stock-feed purposes. Enhancement of genetic and genomic resources for field pea will permit improved understanding of the control of traits relevant to crop productivity and quality. Advances in second-generation sequencing and associated bioinformatics analysis now provide unprecedented opportunities for the development of such resources. The objective of this study was to perform transcriptome sequencing and characterisation from two genotypes of field pea that differ in terms of seed and plant morphological characteristics.
Biswapriya Biswavas Misra's insight:
Background

Field pea (Pisum sativum L.) is a cool-season grain legume that is cultivated world-wide for both human consumption and stock-feed purposes. Enhancement of genetic and genomic resources for field pea will permit improved understanding of the control of traits relevant to crop productivity and quality. Advances in second-generation sequencing and associated bioinformatics analysis now provide unprecedented opportunities for the development of such resources. The objective of this study was to perform transcriptome sequencing and characterisation from two genotypes of field pea that differ in terms of seed and plant morphological characteristics.

Results

Transcriptome sequencing was performed with RNA templates from multiple tissues of the field pea genotypes Kaspa and Parafield. Tissue samples were collected at various growth stages, and a total of 23 cDNA libraries were sequenced using Illumina high-throughput sequencing platforms. A total of 407 and 352 million paired-end reads from the Kaspa and Parafield transcriptomes, respectively were assembled into 129,282 and 149,272 contigs, which were filtered on the basis of known gene annotations, presence of open reading frames (ORFs), reciprocal matches and degree of coverage. Totals of 126,335 contigs from Kaspa and 145,730 from Parafield were subsequently selected as the reference set. Reciprocal sequence analysis revealed that c. 87 % of contigs were expressed in both cultivars, while a small proportion were unique to each genotype. Reads from different libraries were aligned to the genotype-specific assemblies in order to identify and characterise expression of contigs on a tissue-specific basis, of which 87 % were expressed in more than one tissue, while others showed distinct expression patterns in specific tissues, providing unique transcriptome signatures.

Conclusion

This study provided a comprehensive assembled and annotated transcriptome set for field pea that can be used for development of genetic markers, in order to assess genetic diversity, construct linkage maps, perform trait-dissection and implement whole-genome selection strategies in varietal improvement programs, as well to identify target genes for genetic modification approaches on the basis of annotation and expression analysis. In addition, the reference field pea transcriptome will prove highly valuable for comparative genomics studies and construction of a finalised genome sequence.

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The complete mitochondrial genome sequence of the green microalga Lobosphaera (Parietochloris) incisa reveals a new type of palindromic repetit...

Lobosphaera incisa, formerly known as Myrmecia incisa and then Parietochloris incisa, is an oleaginous unicellular green alga belonging to the class Trebouxiophyceae (Chlorophyta). It is the richest known plant source of arachidonic acid, an ω-6 poly-unsaturated fatty acid valued by the pharmaceutical and baby-food industries. It is therefore an organism of high biotechnological interest, and we recently reported the sequence of its chloroplast genome.
Biswapriya Biswavas Misra's insight:
AbstractBackground

Lobosphaera incisa, formerly known as Myrmecia incisa and then Parietochloris incisa, is an oleaginous unicellular green alga belonging to the class Trebouxiophyceae (Chlorophyta). It is the richest known plant source of arachidonic acid, an ω-6 poly-unsaturated fatty acid valued by the pharmaceutical and baby-food industries. It is therefore an organism of high biotechnological interest, and we recently reported the sequence of its chloroplast genome.

Results

We now report the complete sequence of the mitochondrial genome of L. incisa from high-throughput Illumina short-read sequencing. The circular chromosome of 69,997 bp is predicted to encode a total of 64 genes, some harboring specific self-splicing group I and group II introns. Overall, the gene content is highly similar to that of the mitochondrial genomes of other Trebouxiophyceae, with 34 protein-coding, 3 rRNA, and 27 tRNA genes. Genes are distributed in two clusters located on different DNA strands, a bipartite arrangement that suggests expression from two divergent promoters yielding polycistronic primary transcripts. The L. incisa mitochondrial genome contains families of intergenic dispersed DNA repeat sequences that are not shared with other known mitochondrial genomes of Trebouxiophyceae. The most peculiar feature of the genome is a repetitive palindromic repeat, the LIMP (L. Incisa Mitochondrial Palindrome), found 19 times in the genome. It is formed by repetitions of an AACCA pentanucleotide, followed by an invariant 7-nt loop and a complementary repeat of the TGGTT motif. Analysis of the genome sequencing reads indicates that the LIMP can be a substrate for large-scale genomic rearrangements. We speculate that LIMPs can act as origins of replication. Deep sequencing of the L. incisa transcriptome also suggests that the LIMPs with long stems are sites of transcript processing. The genome also contains five copies of a related palindromic repeat, the HyLIMP, with a 10-nt motif related to that of the LIMP.

Conclusions

The mitochondrial genome of L. incisa encodes a unique type of repetitive palindromic repeat sequence, the LIMP, which can mediate genome rearrangements and play a role in mitochondrial gene expression. Experimental studies are needed to confirm and further characterize the functional role(s) of the LIMP.

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