Characterization of the poplar pan-genome by genome-wide identification of structural variation

Many recent studies have emphasized the important role of structural variation (SV) in determining human genetic and phenotypic variation. In plants, studies aimed at elucidating the extent of SV are still in their infancy. Evidence has indicated a high presence and an active role of SV in driving plant genome evolution in different plant species.With the aim of characterizing the size and the composition of the poplar pan-genome, we performed a genome-wide analysis of structural variation in three intercrossable poplar species: Populus nigra, Populus deltoides, and Populus trichocarpa. We detected a total of 7,889 deletions and 10,586 insertions relative to the P. trichocarpa reference genome, covering respectively 33.2?Mb and 62.9?Mb of genomic sequence, and 3,230 genes affected by copy number variation (CNV). The majority of the detected variants are inter-specific in agreement with a recent origin following separation of species.Insertions and deletions (INDELs) were preferentially located in low-gene density regions of the poplar genome and were, for the majority, associated with the activity of transposable elements. Genes affected by SV showed lower-than-average expression levels and higher levels of dN/dS, suggesting that they are subject to relaxed selective pressure or correspond to pseudogenes.Functional annotation of genes affected by INDELs showed over-representation of categories associated with transposable elements activity, while genes affected by genic CNVs showed enrichment in categories related to resistance to stress and pathogens. This study provides a genome-wide catalogue of SV and the first insight on functional and structural properties of the poplar pan-genome

Sequencing of diverse mandarin, pummelo and orange genomes reveals complex history of admixture during citrus domestication

Cultivated citrus are selections from, or hybrids of, wild progenitor species whose identities and contributions to citrus domestication remain controversial. Here we sequence and compare citrus genomes-a high-quality reference haploid clementine genome and mandarin, pummelo, sweet-orange and sour-orange genomes-and show that cultivated types derive from two progenitor species. Although cultivated pummelos represent selections from one progenitor species, Citrus maxima, cultivated mandarins are introgressions of C. maxima into the ancestral mandarin species Citrus reticulata. The most widely cultivated citrus, sweet orange, is the offspring of previously admixed individuals, but sour orange is an F1 hybrid of pure C. maxima and C. reticulata parents, thus implying that wild mandarins were part of the early breeding germplasm. A Chinese wild 'mandarin' diverges substantially from C. reticulata, thus suggesting the possibility of other unrecognized wild citrus species. Understanding citrus phylogeny through genome analysis clarifies taxonomic relationships and facilitates sequence-directed genetic improvement. (Résumé d'auteur

Structural variation and genome complexity: is dispensable really dispensable?

Structural variants (SVs) such as copy number variants (CNVs) and presence/absence variants (PAVs) substantially contribute to genetic variation and have an important effect on phenotypic diversity. Since unbalanced SVs are by definition sequences present only in some individuals, they have therefore been referred to as dispensable genome and are not necessary for survival, even though they may provide an important contribution to phenotypic diversity within the species. However, some multi-copy sequences of the dispensable genomes (e.g., multigene families) may be needed in a given proportion by each individual, thus belonging to a conditionally dispensable portion of the pan-genome. Another interesting aspect reported by recent studies is that the rate at which SVs are formed might be influenced by the mating system and by common environmental stresses. In conclusion the dispensable genome plays an important role in genome evolution and in the complex interplay between the genome and the environment

De novo transcriptome assembly for Pachygrapsus marmoratus, an intertidal brachyuran crab

The marble crab Pachygrapsus marmoratus inhabits the rocky shores of the Mediterranean Sea, Black Sea and East Atlantic Ocean. As other intertidal species, it is considered a model species to study the effects of environmental stressors on natural populations. In this study, we performed Illumina next-generation sequencing on eleven P. marmoratus specimens with the aims to (i) reconstruct their whole transcriptome, (ii) perform a functional annotation of the assembled transcriptome and (iii) develop gene-based markers for future genetic and genomic studies on this as well as other brachyuran species. We obtained a transcriptome assembly constituted by 56,308 unigenes and covering about 60.3 Mbp. We detected 43,915 Simple Sequence Repeats (SSRs) and 192,631 high-quality Single Nucleotide Polymorphisms (SNPs). Due to the scarcity of genomic resources in decapods, and crabs in particular, our results constitute a valuable resource for future studies on brachyuran crabs. The present data also represent a sound resource to investigate biological responses to pollution in intertidal and marine populations

Genomic exploration and molecular marker development in a large and complex conifer genome using RADseq and mRNAseq

We combined restriction site associated DNA sequencing (RADseq) using a hypomethylation-sensitive enzyme and messenger RNA sequencing (mRNAseq) to develop molecular markers for the 16 gigabase genome of[i] Cedrus atlantica[/i], a conifer tree species. With each method, Illumina(®) reads from one individual were used to generate de novo assemblies. SNPs from the RADseq data set were detected in a panel of one single individual and three pools of three individuals each. We developed a flexible script to estimate the ascertainment bias in SNP detection considering the pooling and sampling effects on the probability of not detecting an existing polymorphism. Gene Ontology (GO) and transposable element (TE) search analyses were applied to both data sets. The RADseq and the mRNAseq assemblies represented 0.1% and 0.6% of the genome, respectively. Genome complexity reduction resulted in 17% of the RADseq contigs potentially coding for proteins. This rate was doubled in the mRNAseq data set, suggesting that RADseq also explores noncoding low-repeat regions. The two methods gave very similar GO-slim profiles. As expected, the two assemblies were poor in TE-like sequences (<4% of contigs length). We identified 17,348 single nucleotide polymorphisms (SNPs) in the RADseq data set and 5,714 simple sequence repeats (SSRs) in the transcriptome. A subset of 282 SNPs was validated using the Fluidigm genotyping technology, giving a conversion rate of 50.4%, falling within the expected range for conifers. Increasing sample size had the greatest effect for ascertainment bias reduction. These results validated the utility of the RADseq approach for highly complex genomes such as conifer

Isoprene enhances leaf cytokinin metabolism and induces early senescence.

●Isoprene, a major biogenic volatile hydrocarbon of climate-relevance, indisputably mitigates abiotic stresses in emitting plants. However functional relevance of constitutive isoprene emission in unstressed plants remains contested. Isoprene and cytokinins (CKs) are synthesised from a common substrate and pathway in chloroplasts. It was postulated that isoprene emission may affect CK-metabolism. ●Using transgenic isoprene-emitting (IE) Arabidopsis and isoprene non-emitting (NE) RNAi grey poplars (paired with respective NE and IE genotypes), the life of individual IE and NE leaves from emergence to abscission was followed under stress-free conditions. We monitored plant growth rate, above-ground developmental phenotype, modelled leaf photosynthetic energy status, quantified the abundance of leaf CKs, analyzed Arabidopsis and poplar leaf transcriptomes by RNA-sequencing in presence and absence of isoprene during leaf senescence. ●Isoprene emission by unstressed leaves enhanced the abundance of CKs (isopentenyl adenine and its precursor) by &gt;200%, significantly upregulated genes coding for CK-synthesis, CK-signaling and CK-degradation, hastened plant development, increased chloroplast metabolic rate, altered photosynthetic energy status, induced early leaf senescence in both Arabidopsis and poplar. IE leaves senesced sooner even in decapitated poplars where source-sink relationships and hormone homeostasis were perturbed. ●Constitutive isoprene emission significantly accelerates CK-led leaf and organismal development and induces early senescence independent of growth constraints. Isoprene emission provides an early-riser evolutionary advantage and shortens lifecycle duration to assist rapid diversification in unstressed emitters

Large-scale detection of rare variants via pooled multiplexed next-generation sequencing: towards next-generation Ecotilling.

Common variants, such as those identified by genome-wide association scans, explain only a small proportion of trait variation. Growing evidence suggests that rare functional variants, which are usually missed by genome-wide association scans, play an important role in determining the phenotype. We used pooled multiplexed next-generation sequencing and a customized analysis workflow to detect mutations in five candidate genes for lignin biosynthesis in 768 pooled Populus nigra accessions. We identified a total of 36 non-synonymous single nucleotide polymorphisms, one of which causes a premature stop codon. The most common variant was estimated to be present in 672 of the 1536 tested chromosomes, while the rarest was estimated to occur only once in 1536 chromosomes. Comparison with individual Sanger sequencing in a selected sub-sample confirmed that variants are identified with high sensitivity and specificity, and that the variant frequency was estimated accurately. This proposed method for identification of rare polymorphisms allows accurate detection of variation in many individuals, and is cost-effective compared to individual sequencing

Differential gene expression and chemical patterns of an intertidal crab inhabiting a polluted port and an adjacent marine protected area

The acquisition of data to safeguard marine protected areas located close to ports is important in order to develop plans that allow effective protection from pollution as well as sustainable development of the port. The area Secche della Meloria is a Marine Protected Area (MPA-MEL) three miles from Livorno Harbour (LH), which is characterized by a long history of pollution. Here we studied the bioaccumulation and transcriptomic patterns of the marbled crab, Pachygrapsus marmoratus (Fabricius, 1787) (Crustacea; Brachyura, Grapsidae), inhabiting the two selected sites. Results showed that the two crab populations are significantly different in their chemical composition of trace elements and Polyciclic Aromatic Hydrocarbons (PAHs), and gene expression patterns (1280 DEGs). Enrichment analysis indicated that crabs at LH had the highest stress response genes, and they were associated with higher levels of bioaccumulation detected in body tissues. We are confident that the significant differential gene expression profiles observed between crabs, characterized by significant chemical differences, is associated with responses to contaminant exposure