Adaptor Template Oligo-Mediated Sequencing (ATOM-Seq) is a new ultra-sensitive UMI-based NGS library preparation technology for use with cfDNA and cfRNA.

Liquid biopsy testing utilising Next Generation Sequencing (NGS) is rapidly moving towards clinical adoption for personalised oncology. However, before NGS can fulfil its potential any novel testing approach must identify ways of reducing errors, allowing separation of true low-frequency mutations from procedural artefacts, and be designed to improve upon current technologies. Popular NGS technologies typically utilise two DNA capture approaches; PCR and ligation, which have known limitations and seem to have reached a development plateau with only small, stepwise improvements being made. To maximise the ultimate utility of liquid biopsy testing we have developed a highly versatile approach to NGS: Adaptor Template Oligo Mediated Sequencing (ATOM-Seq). ATOM-Seq's strengths and versatility avoid the major limitations of both PCR- and ligation-based approaches. This technology is ligation free, simple, efficient, flexible, and streamlined, and it offers novel advantages that make it perfectly suited for use on highly challenging clinical material. Using reference and clinical materials, we demonstrate detection of known SNVs down to allele frequencies of 0.1% using as little as 20-25 ng of cfDNA, as well as the ability to detect fusions from RNA. We illustrate ATOM-Seq's suitability for clinical testing by showing high concordance rates between paired cfDNA and FFPE clinical samples.

The evolution of ependymin-related proteins.

BACKGROUND: Ependymins were originally defined as fish-specific secreted glycoproteins involved in central nervous system plasticity and memory formation. Subsequent research revealed that these proteins represent a fish-specific lineage of a larger ependymin-related protein family (EPDRs). EPDRs have now been identified in a number of bilaterian animals and have been implicated in diverse non-neural functions. The recent discoveries of putative EPDRs in unicellular holozoans and an expanded EPDR family with potential roles in conspecific communication in crown-of-thorns starfish suggest that the distribution and diversity of EPDRs is significantly broader than currently understood. RESULTS: We undertook a systematic survey to determine the distribution and evolution of EPDRs in eukaryotes. In addition to Bilateria, EPDR genes were identified in Cnidaria, Placozoa, Porifera, Choanoflagellatea, Filasterea, Apusozoa, Amoebozoa, Charophyta and Percolozoa, and tentatively in Cercozoa and the orphan group Malawimonadidae. EPDRs appear to be absent from prokaryotes and many eukaryote groups including ecdysozoans, fungi, stramenopiles, alveolates, haptistans and cryptistans. The EPDR family can be divided into two major clades and has undergone lineage-specific expansions in a number of metazoan lineages, including in poriferans, molluscs and cephalochordates. Variation in a core set of conserved residues in EPDRs reveals the presence of three distinct protein types; however, 3D modelling predicts overall protein structures to be similar. CONCLUSIONS: Our results reveal an early eukaryotic origin of the EPDR gene family and a dynamic pattern of gene duplication and gene loss in animals. This research provides a phylogenetic framework for the analysis of the functional evolution of this gene family.

Amphioxus Sp5 is a member of a conserved Specificity Protein complement and is modulated by Wnt/ß-catenin signalling.

A cluster of three Specificity Protein (Sp) genes (Sp1-4, Sp5 and Sp6-9) is thought to be ancestral in both chordates and the wider Eumetazoa. Sp5 and Sp6-9 gene groups are associated with embryonic growth zones, such as tailbuds, and are both Wnt/ß-catenin signalling pathway members and targets. Currently, there are conflicting reports as to the number and identity of Sp genes in the cephalochordates, the sister group to the vertebrates and urochordates. We confirm the SP complement of Branchiostoma belcheri and Branchiostoma lanceolatum, as well as their genomic arrangement, protein domain structure and residue frequency. We assay Sp5 expression in B. lanceolatum embryos, and determine its response to pharmacologically increased ß-catenin signalling. Branchiostoma possesses three Sp genes, located on the same genomic scaffold. Phylogenetic and domain structure analyses are consistent with their identification as SP1-4, SP5 and SP6-9, although SP1-4 contains a novel glutamine-rich N-terminal region. SP5 is expressed in axial mesoderm and neurectoderm, and marks the cerebral vesicle and presumptive pharynx. Early exposure to increased ß-catenin caused ubiquitous SP5 expression in late gastrula, while later treatment at gastrula stages reduced SP5 expression in the posterior growth zone during axis elongation. Amphioxus possess a typical invertebrate eumetazoan SP complement, and SP5 expression in embryos is well conserved with vertebrate homologues. Its expression in the tailbud, a posterior growth zone, is consistent with expression seen in other bilaterians. Branchiostoma SP5 shows a dynamic response to Wnt/ß-catenin signalling.