Comparative Hessian Fly Larval Transcriptomics Provides Novel Insight into Host and Nonhost Resistance

The Hessian fly is a destructive pest of wheat. Employing additional molecular strategies can complement wheat’s native insect resistance. However, this requires functional characterization of Hessian-fly-responsive genes, which is challenging because of wheat genome complexity. The diploid Brachypodium distachyon (Bd) exhibits nonhost resistance to Hessian fly and displays phenotypic/molecular responses intermediate between resistant and susceptible host wheat, offering a surrogate genome for gene characterization. Here, we compared the transcriptomes of Biotype L larvae residing on resistant/susceptible wheat, and nonhost Bd plants. Larvae from susceptible wheat and nonhost Bd plants revealed similar molecular responses that were distinct from avirulent larval responses on resistant wheat. Secreted salivary gland proteins were strongly up-regulated in all larvae. Genes from various biological pathways and molecular processes were up-regulated in larvae from both susceptible wheat and nonhost Bd plants. However, Bd larval expression levels were intermediate between larvae from susceptible and resistant wheat. Most genes were down-regulated or unchanged in avirulent larvae, correlating with their inability to establish feeding sites and dying within 4–5 days after egg-hatch. Decreased gene expression in Bd larvae, compared to ones on susceptible wheat, potentially led to developmentally delayed 2nd-instars, followed by eventually succumbing to nonhost resistance defense mechanisms.

Application of Genomics Approaches for the Improvement in Ascochyta Blight Resistance in Chickpea

Advancements in high-throughput genotyping and sequencing technologies are enabling the development of a vast range of genomic tools and resources for a new revolution in plant breeding. Several genotyping-by-sequencing (GBS) methods including capture-based, genome complexity reduction and sequencing of cDNA (GBS-t) are available for application in trait dissection, association mapping, and genomic selection (GS) in crop plants. The aims of this study were to identify genomic regions conferring resistance to Ascochyta blight (AB) introgressed from the wild Cicer echinospernum into the domesticated C. arietinum, through a conventional recombinant inbred population genotyped using a variety of GBS methods. Evaluation of GBS methods revealed that capture-based approaches are robust and reproducible while GBS-t is rapid and flexible. A genetic linkage map consisting of 5886 polymorphic loci spanning 717.26 cM was generated. Using field phenotyping data from two years, a single genomic region on LG4 was identified with quantitative trait loci (QTL) mapping. Both GBS methods reported in this study are well suited for applications in genomics assisted plant breeding. Linked markers for AB resistance, identified in the current study, provide an important resource for the deployment into chickpea breeding programs for marker-assisted selection (MAS).

Viromics unveils extraordinary genetic diversity of the family Closteroviridae in wild citrus

Our knowledge of citrus viruses is largely skewed toward virus pathology in cultivated orchards. Little is known about the virus diversity in wild citrus species. Here, we used a metatranscriptomics approach to characterize the virus diversity in a wild citrus habitat within the proposed center of the origin of citrus plants. We discovered a total of 44 virus isolates that could be classified into species Citrus tristeza virus and putative species citrus associated ampelovirus 1, citrus associated ampelovirus 2, and citrus virus B within the family Closteroviridae, providing important information to explore the factors facilitating outbreaks of citrus viruses and the evolutionary history of the family Closteroviridae. We found that frequent horizontal gene transfer, gene duplication, and alteration of expression strategy have shaped the genome complexity and diversification of the family Closteroviridae. Recombination frequently occurred among distinct Closteroviridae members, thereby facilitating the evolution of Closteroviridae. Given the potential emergence of similar wild-citrus-originated novel viruses as pathogens, the need for surveillance of their pathogenic and epidemiological characteristics is of utmost priority for global citrus production.

Sequencing an F1 hybrid of Silurus asotus and S. meridionalis enabled the assembly of high-quality parental genomes

Genome complexity such as heterozygosity may heavily influence its de novo assembly. Sequencing somatic cells of the F1 hybrids harboring two sets of genetic materials from both of the paternal and maternal species may avoid alleles discrimination during assembly. However, the feasibility of this strategy needs further assessments. We sequenced and assembled the genome of an F1 hybrid between Silurus asotus and S. meridionalis using the SequelII platform and Hi-C scaffolding technologies. More than 300 Gb raw data were generated, and the final assembly obtained 2344 scaffolds composed of 3017 contigs. The N50 length of scaffolds and contigs was 28.55 Mb and 7.49 Mb, respectively. Based on the mapping results of short reads generated for the paternal and maternal species, each of the 29 chromosomes originating from S. asotus and S. meridionalis was recognized. We recovered nearly 94% and 96% of the total length of S. asotus and S. meridionalis. BUSCO assessments and mapping analyses suggested that both genomes had high completeness and accuracy. Further analyses demonstrated the high collinearity between S. asotus, S. meridionalis, and the related Pelteobagrus fulvidraco. Comparison of the two genomes with that assembled only using the short reads from non-hybrid parental species detected a small portion of sequences that may be incorrectly assigned to the different species. We supposed that at least part of these situations may have resulted from mitotic recombination. The strategy of sequencing the F1 hybrid genome can recover the vast majority of the parental genomes and may improve the assembly of complex genomes.

Evolution of default genetic control mechanisms

We present a model of the evolution of control systems in a genome under environmental constraints. The model conceptually follows the Jacob and Monod model of gene control. Genes contain control elements which respond to the internal state of the cell as well as the environment to control expression of a coding region. Control and coding regions evolve to maximize a fitness function between expressed coding sequences and the environment. The model was run 118 times to an average of 1.4∙106 ‘generations’ each with a range of starting parameters probed the conditions under which genomes evolved a ‘default style’ of control. Unexpectedly, the control logic that evolved was not significantly correlated to the complexity of the environment. Genetic logic was strongly correlated with genome complexity and with the fraction of genes active in the cell at any one time. More complex genomes correlated with the evolution of genetic controls in which genes were active (‘default on’), and a low fraction of genes being expressed correlated with a genetic logic in which genes were biased to being inactive unless positively activated (‘default off’ logic). We discuss how this might relate to the evolution of the complex eukaryotic genome, which operates in a ‘default off’ mode.

Histone Sample Preparation for Bottom-Up Mass Spectrometry: A Roadmap to Informed Decisions

Histone-based chromatin organization enabled eukaryotic genome complexity. This epigenetic control mechanism allowed for the differentiation of stable gene-expression and thus the very existence of multicellular organisms. This existential role in biology makes histones one of the most complexly modified molecules in the biotic world, which makes these key regulators notoriously hard to analyze. We here provide a roadmap to enable fast and informed selection of a bottom-up mass spectrometry sample preparation protocol that matches a specific research question. We therefore propose a two-step assessment procedure: (i) visualization of the coverage that is attained for a given workflow and (ii) direct alignment between runs to assess potential pitfalls at the ion level. To illustrate the applicability, we compare four different sample preparation protocols while adding a new enzyme to the toolbox, i.e., RgpB (GingisREX®, Genovis, Lund, Sweden), an endoproteinase that selectively and efficiently cleaves at the c-terminal end of arginine residues. Raw data are available via ProteomeXchange with identifier PXD024423.

Next Generation Sequencing Technology in the Clinic and Its Challenges

Data analysis has become a crucial aspect in clinical oncology to interpret output from next-generation sequencing-based testing. NGS being able to resolve billions of sequencing reactions in a few days has consequently increased the demand for tools to handle and analyze such large data sets. Many tools have been developed since the advent of NGS, featuring their own peculiarities. Increased awareness when interpreting alterations in the genome is therefore of utmost importance, as the same data using different tools can provide diverse outcomes. Hence, it is crucial to evaluate and validate bioinformatic pipelines in clinical settings. Moreover, personalized medicine implies treatment targeting efficacy of biological drugs for specific genomic alterations. Here, we focused on different sequencing technologies, features underlying the genome complexity, and bioinformatic tools that can impact the final annotation. Additionally, we discuss the clinical demand and design for implementing NGS.

Fine-tuning the performance of ddRAD-seq in the peach genome

The advance of Next Generation Sequencing (NGS) technologies allows high-throughput genotyping at a reasonable cost, although, in the case of peach, this technology has been scarcely developed. To date, only a standard Genotyping by Sequencing approach (GBS), based on a single restriction with ApeKI to reduce genome complexity, has been applied in peach. In this work, we assessed the performance of the double-digest RADseq approach (ddRADseq), by testing 6 double restrictions with the restriction profile generated with ApeKI. The enzyme pair PstI/MboI retained the highest number of loci in concordance with the in silico analysis. Under this condition, the analysis of a diverse germplasm collection (191 peach genotypes) yielded 200,759,000 paired-end (2 × 250 bp) reads that allowed the identification of 113,411 SNP, 13,661 InDel and 2133 SSR. We take advantage of a wide sample set to describe technical scope of the platform. The novel platform presented here represents a useful tool for genomic-based breeding for peach.

Histone sample preparation for bottom-up mass spectrometry: a roadmap to informed decisions

Histone-based chromatin organization enabled eukaryotic genome complexity. This epigenetic control mechanism allowed for the differentiation of stable gene-expression and thus the very existence of multicellular organisms. This existential role in biology makes histones one of the most complexly modified molecules in the biotic world, which makes these key regulators notoriously hard to analyze. We here provide a roadmap to enable fast and informed selection of a bottom-up mass spectrometry sample preparation protocol that matches a specific research question. We therefore propose a two-step assessment procedure: (i) visualization of the coverage that is attained for a given workflow and (ii) direct alignment-between-runs to assess potential pitfalls at the ion level. To illustrate the applicability, we compare four different sample preparation protocols, while adding a new enzyme to the toolbox, i.e., RgpB (GingisREX, Genovis), an endoproteinase that selectively and efficiently cleaves at the c-terminal end of arginine residues.