Genome‐wide characterization and expression analysis of genetic variants in sweet orange

Heterozyosity is an important feature of many plant genomes, and is related to heterosis. Sweet orange, a highly heterozygous species, is thought to have originated from an inter‐species hybrid between pummelo and mandarin. To investigate the heterozygosity of the sweet orange genome and examine how this heterozygosity affects gene expression, we characterized the genome of Valencia orange for single nucleotide variations (SNVs), small insertions and deletions (InDels) and structural variations (SVs), and determined their functional effects on protein‐coding genes and non‐coding sequences. Almost half of the genes containing large‐effect SNVs and InDels were expressed in a tissue‐specific manner. We identified 3542 large SVs (>50 bp), including deletions, insertions and inversions. Most of the 296 genes located in large‐deletion regions showed low expression levels. RNA‐Seq reads and DNA sequencing reads revealed that the alleles of 1062 genes were differentially expressed. In addition, we detected approximately 42 Mb of contigs that were not found in the reference genome of a haploid sweet orange by de novo assembly of unmapped reads, and annotated 134 protein‐coding genes within these contigs. We discuss how this heterozygosity affects the quality of genome assembly. This study advances our understanding of the genome architecture of sweet orange, and provides a global view of gene expression at heterozygous loci.     

De novo genome assembly of the stress tolerant forest species Casuarina equisetifolia provides insight into secondary growth

Casuarina equisetifolia (C. equisetifolia), a conifer‐like angiosperm with resistance to typhoon and stress tolerance, is mainly cultivated in the coastal areas of Australasia. C. equisetifolia, making it a valuable model to study secondary growth associated genes and stress‐tolerance traits. However, the genome sequence is unavailable and therefore wood‐associated growth rate and stress resistance at the molecular level is largely unexplored. We therefore constructed a high‐quality draft genome sequence of C. equisetifolia by a combination of Illumina second‐generation sequencing reads and Pacific Biosciences single‐molecule real‐time (SMRT) long reads to advance the investigation of this species. Here, we report the genome assembly, which contains approximately 300 megabases (Mb) and scaffold size of N50 is 1.06 Mb. Additionally, gene annotation, assisted by a combination of prediction and RNA‐seq data, generated 29 827 annotated protein‐coding genes and 1983 non‐coding genes, respectively. Furthermore, we found that the total number of repetitive sequences account for one‐third of the genome assembly. Here we also construct the genome‐wide map of DNA modification, such as two novel forms N⁶‐adenine (6mA) and N4‐methylcytosine (4mC) at the level of single‐nucleotide resolution using single‐molecule real‐time (SMRT) sequencing. Interestingly, we found that 17% of 6mA modification genes and 15% of 4mC modification genes also included alternative splicing events. Finally, we investigated cellulose, hemicellulose, and lignin‐related genes, which were associated with secondary growth and contained different DNA modifications. The high‐quality genome sequence and annotation of C. equisetifolia in this study provide a valuable resource to strengthen our understanding of the diverse traits of trees.     

In Vino Veritas: An Invitation for Ambitious,Collaborative Proteogenomics Campaigns on Plant and Animal Models

Vitis vinifera has been an emblematic plant for humans since the Neolithic period. Human civilization has been shaped by its domestication as both its medicinal and nutritional values were exploited. It is now cultivated on all habitable continents, and more than 5000 varieties have been developed. A global passion for the art of wine fuels innovation and a profound desire for knowledge on this plant. The genome sequence of a homozygotic cultivar and several RNA‐seq datasets on other varieties have been released paving the way to gaining further insight into its biology and tailoring improvements to varieties. However, its genome annotation remains unpolished. In this issue of Proteomics, Chapman and Bellgard (Proteomics 2017, 17, 1700197) discuss how proteogenomics can help improve genome annotation. By mining shotgun proteomics data, they defined new protein‐coding genes, refined gene structures, and corrected numerous mRNA splicing events. This stimulating study shows how large international consortia could work together to improve plant and animal genome annotation on a large scale. To achieve this aim, time should be invested to generate comprehensive, high‐quality experimental datasets for a wide range of well‐defined lineages and exploit them with pipelines capable of handling giant datasets.     

A large‐scale proteogenomics study of apicomplexan pathogens—Toxoplasma gondii and Neospora caninum

Proteomics data can supplement genome annotation efforts, for example being used to confirm gene models or correct gene annotation errors. Here, we present a large‐scale proteogenomics study of two important apicomplexan pathogens: Toxoplasma gondii and Neospora caninum. We queried proteomics data against a panel of official and alternate gene models generated directly from RNASeq data, using several newly generated and some previously published MS datasets for this meta‐analysis. We identified a total of 201 996 and 39 953 peptide‐spectrum matches for T. gondii and N. caninum, respectively, at a 1% peptide FDR threshold. This equated to the identification of 30 494 distinct peptide sequences and 2921 proteins (matches to official gene models) for T. gondii, and 8911 peptides/1273 proteins for N. caninum following stringent protein‐level thresholding. We have also identified 289 and 140 loci for T. gondii and N. caninum, respectively, which mapped to RNA‐Seq‐derived gene models used in our analysis and apparently absent from the official annotation (release 10 from EuPathDB) of these species. We present several examples in our study where the RNA‐Seq evidence can help in correction of the current gene model and can help in discovery of potential new genes. The findings of this study have been integrated into the EuPathDB. The data have been deposited to the ProteomeXchange with identifiers PXD000297and PXD000298.