The recent advent of long read sequencing technologies, such as Pacific Biosciences (PacBio) and Oxford Nanopore technology (ONT), have led to substantial improvements in accuracy and computational cost in sequencing genomes. However, de novo whole-genome assembly still presents significant challenges related to the quality of the results. Pursuing de novo whole-genome assembly remains a formidable challenge, underscored by intricate considerations surrounding computational demands and result quality. As sequencing accuracy and throughput steadily advance, a continuous stream of innovative assembly tools floods the field. Navigating this dynamic landscape necessitates a reasonable choice of sequencing platform, depth, and assembly tools to orchestrate high-quality genome reconstructions. This comprehensive review delves into the intricate interplay between cutting-edge long read sequencing technologies, assembly methodologies, and the ever-evolving field of genomics. With a focus on addressing the pivotal challenges and harnessing the opportunities presented by these advancements, we provide an in-depth exploration of the crucial factors influencing the selection of optimal strategies for achieving robust and insightful genome assemblies.

Maritime pine (Pinus pinasterAit.) is a widely distributed conifer species in Southwestern Europe and one of the most advanced models for conifer research. In the current work, comprehensive characterization of the maritime pine transcriptome was performed using a combination of two different next‐generation sequencing platforms, 454 and Illumina. De novo assembly of the transcriptome provided a catalogue of 26 020 unique transcripts in maritime pine trees and a collection of 9641 full‐length cDNAs. Quality of the transcriptome assembly was validated by RT‐PCR amplification of selected transcripts for structural and regulatory genes. Transcription factors and enzyme‐encoding transcripts were annotated. Furthermore, the available sequencing data permitted the identification of polymorphisms and the establishment of robust single nucleotide polymorphism (SNP) and simple‐sequence repeat (SSR) databases for genotyping applications and integration of translational genomics in maritime pine breeding programmes. All our data are freely available at SustainpineDB, the P. pinaster expressional database. Results reported here on the maritime pine transcriptome represent a valuable resource for future basic and applied studies on this ecological and economically important pine species.

In spite of the biological and economic importance of plants, relatively few plant species have been sequenced. Only the genome sequence of plants with relatively small genomes, most of them angiosperms, in particular eudicots, has been determined. The arrival of next-generation sequencing technologies has allowed the rapid and efficient development of new genomic resources for non-model or orphan plant species. But the sequencing pace of plants is far from that of animals and microorganisms. This review focuses on the typical challenges of plant genomes that can explain why plant genomics is less developed than animal genomics. Explanations about the impact of some confounding factors emerging from the nature of plant genomes are given. As a result of these challenges and confounding factors, the correct assembly and annotation of plant genomes is hindered, genome drafts are produced, and advances in plant genomics are delayed.

Ripening is a complex physiological process that involves changes in reactive nitrogen and oxygen species that govern the shelf-life and quality of fruits. Nitric oxide (NO)-dependent changes in the sweet pepper fruit transcriptome were determined by treating fruits at the initial breaking point stage with NO gas. Fruits were also harvested at the immature (green) and ripe (red) stages. Fruit ripening in the absence of NO resulted in changes in the abundance of 8805 transcripts whose function could be identified. Among these, functional clusters associated with reactive oxygen/nitrogen species and lipid metabolism were significantly modified. NO treatment resulted in the differential expression of 498 genes framed within these functional categories. Biochemical analysis revealed that NO treatment resulted in changes in fatty acid profiling, glutathione and proline contents, and the extent of lipid peroxidation, as well as increases in the activity of ascorbate peroxidase and lipoxygenase. These data provide supporting evidence for the crucial role of NO in the ripening of pepper fruit.

Senegalese sole (Solea senegalensis) and common sole (S. solea) are two economically and evolutionary important flatfish species both in fisheries and aquaculture. Although some genomic resources and tools were recently described in these species, further sequencing efforts are required to establish a complete transcriptome, and to identify new molecular markers. Moreover, the comparative analysis of transcriptomes will be useful to understand flatfish evolution. A comprehensive characterization of the transcriptome for each species was carried out using a large set of Illumina data (more than 1,800 millions reads for each sole species) and 454 reads (more than 5 millions reads only in S. senegalensis), providing coverages ranging from 1,384x to 2,543x. After a de novo assembly, 45,063 and 38,402 different transcripts were obtained, comprising 18,738 and 22,683 full-length cDNAs in S. senegalensis and S. solea, respectively. A reference transcriptome with the longest unique transcripts and putative non-redundant new transcripts was established for each species. A subset of 11,953 reference transcripts was qualified as highly reliable orthologs (>97% identity) between both species. A small subset of putative species-specific, lineage-specific and flatfish-specific transcripts were also identified. Furthermore, transcriptome data permitted the identification of single nucleotide polymorphisms and simple-sequence repeats confirmed by FISH to be used in further genetic and expression studies. Moreover, evidences on the retention of crystallins crybb1, crybb1-like and crybb3 in the two species of soles are also presented. Transcriptome information was applied to the design of a microarray tool in S. senegalensis that was successfully tested and validated by qPCR. Finally, transcriptomic data were hosted and structured at SoleaDB. Transcriptomes and molecular markers identified in this study represent a valuable source for future genomic studies in these economically important species. Orthology analysis provided new clues regarding sole genome evolution indicating a divergent evolution of crystallins in flatfish. The design of a microarray and establishment of a reference transcriptome will be useful for large-scale gene expression studies. Moreover, the integration of transcriptomic data in the SoleaDB will facilitate the management of genomic information in these important species.

Nowadays, Huanglongbing (HLB) disease, associated with Candidatus Liberibacter asiaticus (CLas), seriously affects citriculture worldwide, and no cure is currently available. Transcriptomic analysis of host–pathogen interaction is the first step to understand the molecular landscape of a disease. Previous works have reported the transcriptome profiling in response to HLB in different susceptible citrus species; however, similar studies in tolerant citrus species, including Mexican lime, are limited. In this work, we have obtained an RNA-seq-based differential expression profile of Mexican lime plants challenged against CLas infection, at both asymptomatic and symptomatic stages. Typical HLB-responsive differentially expressed genes (DEGs) are involved in photosynthesis, secondary metabolism, and phytohormone homeostasis. Enrichment of DEGs associated with biotic response showed that genes related to cell wall, secondary metabolism, transcription factors, signaling, and redox reactions could play a role in the tolerance of Mexican lime against CLas infection. Interestingly, despite some concordance observed between transcriptional responses of different tolerant citrus species, a subset of DEGs appeared to be species-specific. Our data highlights the importance of studying the host response during HLB disease using as model tolerant citrus species, in order to design new and opportune diagnostic and management methods.