Clinical and genetic diagnosis of thirteen Japanese patients with hereditary spherocytosis

Hereditary spherocytosis is the most frequent cause of hereditary hemolytic anemia and is classified into five subtypes (SPH1-5) according to OMIM. Because the clinical and laboratory features of patients with SPH1-5 are variable, it is difficult to classify these patients into the five subtypes based only on these features. We performed target capture sequencing in 51 patients with hemolytic anemia associated with/without morphological abnormalities in red blood cells. Thirteen variants were identified in five hereditary spherocytosis-related genes (six in ANK1 [SPH1]; four in SPTB [SPH2]; and one in each of SPTA1 [SPH3], SLC4A1 [SPH4], and EPB42 [SPH5]). Among these variants, seven were novel. The distribution pattern of the variants was different from that reported previously in Japan but similar to those reported in other Asian countries. Comprehensive genomic analysis would be useful and recommended, especially for patients without a detailed family history and those receiving frequent blood transfusions due to chronic hemolytic anemia.

A target capture approach for phylogenomic analyses at multiple evolutionary timescales in rosewoods (Dalbergia spp.) and the legume family (Fabaceae)

Understanding the genetic changes associated with the evolution of biological diversity is of fundamental interest to molecular ecologists. The assessment of genetic variation at hundreds or thousands of unlinked genetic loci forms a sound basis to address questions ranging from micro- to macro-evolutionary timescales, and is now possible thanks to advances in sequencing technology. Major difficulties are associated with i) the lack of genomic resources for many taxa, especially from tropical biodiversity hotspots, ii) scaling the numbers of individuals analyzed and loci sequenced, and iii) building tools for reproducible bioinformatic analyses of such datasets. To address these challenges, we developed a set of target capture probes for phylogenomic studies of the highly diverse, pantropically distributed and economically significant rosewoods (Dalbergia spp.), explored the performance of an overlapping probe set for target capture across the legume family (Fabaceae), and built a general-purpose bioinformatics pipeline. Phylogenomic analyses of Dalbergia species from Madagascar yielded highly resolved and well supported hypotheses of evolutionary relationships. Population genomic analyses identified differences between closely related species and revealed the existence of a potentially new species, suggesting that the diversity of Malagasy Dalbergia species has been underestimated. Analyses at the family level corroborated previous findings by the recovery of monophyletic subfamilies and many well-known clades, as well as high levels of gene tree discordance, especially near the root of the family. The new genomic and bioinformatics resources will hopefully advance systematics and ecological genetics research in legumes, and promote conservation of the highly diverse and endangered Dalbergia rosewoods.

A target capture approach for phylogenomic analyses at multiple evolutionary timescales in rosewoods (Dalbergia spp.) and the legume family (Fabaceae)

Understanding the genetic changes associated with the evolution of biological diversity is of fundamental interest to molecular ecologists. The assessment of genetic variation at hundreds or thousands of unlinked genetic loci forms a sound basis to address questions ranging from micro- to macro-evolutionary timescales, and is now possible thanks to advances in sequencing technology. Major difficulties are associated with i) the lack of genomic resources for many taxa, especially from tropical biodiversity hotspots, ii) scaling the numbers of individuals analyzed and loci sequenced, and iii) building tools for reproducible bioinformatic analyses of such datasets. To address these challenges, we developed a set of target capture probes for phylogenomic studies of the highly diverse, pantropically distributed and economically significant rosewoods (Dalbergia spp.), explored the performance of an overlapping probe set for target capture across the legume family (Fabaceae), and built a general-purpose bioinformatics pipeline. Phylogenomic analyses of Dalbergia species from Madagascar yielded highly resolved and well supported hypotheses of evolutionary relationships. Population genomic analyses identified differences between closely related species and revealed the existence of a potentially new species, suggesting that the diversity of Malagasy Dalbergia species has been underestimated. Analyses at the family level corroborated previous findings by the recovery of monophyletic subfamilies and many well-known clades, as well as high levels of gene tree discordance, especially near the root of the family. The new genomic and bioinformatics resources will hopefully advance systematics and ecological genetics research in legumes, and promote conservation of the highly diverse and endangered Dalbergia rosewoods.

A customised target capture sequencing tool for molecular identification of Aloe vera and relatives

Plant molecular identification studies have, until recently, been limited to the use of highly conserved markers from plastid and other organellar genomes, compromising resolution in highly diverse plant clades. Due to their higher evolutionary rates and reduced paralogy, low-copy nuclear genes overcome this limitation but are difficult to sequence with conventional methods and require high-quality input DNA. Aloe vera and its relatives in the Alooideae clade (Asphodelaceae, subfamily Asphodeloideae) are of economic interest for food and health products and have horticultural value. However, pressing conservation issues are increasing the need for a molecular identification tool to regulate the trade. With > 600 species and an origin of ± 15 million years ago, this predominantly African succulent plant clade is a diverse and taxonomically complex group for which low-copy nuclear genes would be desirable for accurate species discrimination. Unfortunately, with an average genome size of 16.76 pg, obtaining high coverage sequencing data for these genes would be prohibitively costly and computationally demanding. We used newly generated transcriptome data to design a customised RNA-bait panel targeting 189 low-copy nuclear genes in Alooideae. We demonstrate its efficacy in obtaining high-coverage sequence data for the target loci on Illumina sequencing platforms, including degraded DNA samples from museum specimens, with considerably improved phylogenetic resolution. This customised target capture sequencing protocol has the potential to confidently indicate phylogenetic relationships of Aloe vera and related species, as well as aid molecular identification applications.

GSA: an independent development algorithm for calling copy number and detecting homologous recombination deficiency (HRD) from target capture sequencing

Background The gain or loss of large chromosomal regions or even whole chromosomes is termed as genomic scarring and can be observed as copy number variations resulting from the failure of DNA damage repair. Results In this study, a new algorithm called genomic scar analysis (GSA) has developed and validated to calculate homologous recombination deficiency (HRD) score. The two critical submodules were tree recursion (TR) segmentation and filtering, and the estimation and correction of the tumor purity and ploidy. Then, this study evaluated the rationality of segmentation and genotype identification by the GSA algorithm and compared with other two algorithms, PureCN and ASCAT, found that the segmentation result of GSA algorithm was more logical. In addition, the results indicated that the GSA algorithm had an excellent predictive effect on tumor purity and ploidy, if the tumor purity was more than 20%. Furtherly, this study evaluated the HRD scores and BRCA1/2 deficiency status of 195 clinical samples, and the results indicated that the accuracy was 0.98 (comparing with Affymetrix OncoScan™ assay) and the sensitivity was 95.2% (comparing with BRCA1/2 deficiency status), both were well-behaved. Finally, HRD scores and 16 genes mutations (TP53 and 15 HRR pathway genes) were analyzed in 17 cell lines, the results showed that there was higher frequency in HRR pathway genes in high HRD score samples. Conclusions This new algorithm, named as GSA, could effectively and accurately calculate the purity and ploidy of tumor samples through NGS data, and then reflect the degree of genomic instability and large-scale copy number variations of tumor samples.

Decoding the olfactory map: targeted transcriptomics link olfactory receptors to glomeruli

Sensory processing in olfactory systems is organized across olfactory bulb glomeruli, wherein axons of peripheral sensory neurons expressing the same olfactory receptor co-terminate to transmit receptor-specific activity to central neurons. Understanding how receptors map to glomeruli is therefore critical to understanding olfaction. High-throughput spatial transcriptomics is a rapidly advancing field, but low-abundance olfactory receptor expression within glomeruli has previously precluded high-throughput mapping of receptors to glomeruli. Here we combined sequential sectioning along the anteroposterior, dorsoventral, and mediolateral axes with target capture enrichment sequencing to overcome low-abundance target expression. This strategy allowed us to spatially map 86% of olfactory receptors across the olfactory bulb and uncover a relationship between OR sequence and glomerular position.