The use of morphological, developmental, and plant nitrogen traits in a selection scheme in soybean

A three year study was conducted in which the objective was to determine the response in seed yield from a tandem selection scheme in which the first step was to select desirable lines from each of two single cross populations for morphological and developmental traits, using independent culling. The second step was to select for high plant nitrogen (N) content at the R5 developmental stage. The genetic material was 213 F6 derived indeterminate lines from three maturity groups (79 early, 93 medium, and 41 late lines) from a single cross population (IX139) and 102 lines from three different stem termination types (28 determinate, 40 semideterminate, and 34 indeterminate lines) from a different cross (IX149);Yield increase from the first part of the selection scheme in the early, medium, and late maturity groups of IX139 was 2.2, 1.9, and 5.6%, respectively with a mean of 3.2% across locations and years. Selection for morphological and developmental traits in the determinate group of IX149 decreased the seed yield by 4.6% and increased the seed yield in the semideterminate and indeterminate groups of IXl49 by 1.1 and 4.9% across locations and years. In the second portion of the tandem selection scheme, selection for high plant N content at R5 decreased the seed yield by an average of 0.3% across the maturity groups of IX139 and increased the yield by 1.4% across the three stem termination types of IX149;In assessing the overall response to tandem selection in all groups within IX139 and IX149, most groups responded positively, but the determinates decreased in seed yield by 4.1%;Plant nitrogen content at the R5 stage, because of its low association with seed yield and the inconsistent seed yield response to selection does not appear to play a significant role in determining the final yield

Fabrication method, microstructural characteristics, and hardness behavior of an interpenetrating phases hybrid aluminum/alumina-nanodiamond composite

In the present work, the addition effects of nanodiamond (ND) on the microstructure and hardness behavior of interpenetrating phases hybrid Al/Al2O3 metal matrix composites were investigated. The fabrication of the composites was done via a two-step process. In the first step, hybrid Al2O3-ND preforms were prepared, and then molten pure Al alloy was infiltrated into the preforms. The preforms were fabricated by the replica method using a polyurethane foam and an Al2O3-ND slurry with various ND contents (0, 1, 3, and 10 vol%). The preforms were sintered at 1500 °C for 4 h under argon gas protection. Finally, the composites were fabricated by Al melt infiltration into the preforms via the squeeze casting method. The microstructure of the fabricated composites was analyzed using optical and scanning electron microscopes. The hardness of the composites was measured using a Vickers hardness tester. The results of the microstructural evaluations demonstrated a good distribution of ND in the preform. By increasing the ND content from 0 to 10 vol%, the matrix average grain size decreased from 143 μm to 76 μm. The results of the Vickers hardness test showed that increasing the volume percentage of ND increased the composite hardness to 263.8 Vickers at 10 vol%. The two main strengthening mechanisms for these composites are the Orowwn mechanism (volume fraction of ND particles) and the Hal-Petch mechanism (grain size), which affect the hardness behavior

Dystrophic Epidermolysis Bullosa: COL7A1 Mutation Landscape in a Multi-Ethnic Cohort of 152 Extended Families with High Degree of Customary Consanguineous Marriages

Dystrophic epidermolysis bullosa is a heritable skin disease manifesting with sub-lamina densa blistering, erosions, and chronic ulcers. COL7A1, encoding type VII collagen, has been identified as the candidate gene for dystrophic epidermolysis bullosa. In this study, we have identified COL7A1 mutations in a large multi-ethnic cohort of 152 extended Iranian families with high degree of consanguinity. The patients were diagnosed by clinical manifestations, histopathology, and immunoepitope mapping. Mutation detection consisted of a combination of single nucleotide polymorphism-based whole-genome homozygosity mapping, Sanger sequencing, and gene-targeted next-generation sequencing. A total of 104 distinct mutations in COL7A1 were identified in 149 of 152 families (98%), 56 (53%) of them being previously unreported. Ninety percent of these mutations were homozygous recessive, reflecting consanguinity in these families. Three recurrent mutations were identified in five or more families, and haplotype analysis suggested a founder effect in two of them. In conclusion, COL7A1 harbored mutations in the overwhelming majority of patients with dystrophic epi-dermolysis bullosa, and most of them in this Iranian cohort were consistent with autosomal recessive inheri-tance. The mutation profile attests to the impact of consanguinity in these families

Gene-Targeted Next Generation Sequencing Identifies PNPLA1 Mutations in Patients with a Phenotypic Spectrum of Autosomal Recessive Congenital Ichthyosis: The Impact of Consanguinity

Heritable forms of ichthyoses, also referred to as generalized Mendelian disorders of cornification, are phenotypically a highly heterogeneous group of conditions caused by mutations in a number of genes playing a role in keratinocyte differentiation and epidermal barrier function (Baden and Digiovanna, 2013; Schmuth et al., 2013). These diseases are characterized by scaling and hyperkeratosis with associated cutaneous and extracutaneous features. This group of disorders is also genetically heterogeneous, with autosomal dominant, autosomal recessive, and X-linked inheritance being described. A specific subgroup of inherited ichthyoses is the autosomal recessive congenital ichthyosis (ARCI), with many newborns presenting as collodion babies, but the subsequent clinical presentation and the spectrum of severity can be highly variable (Richard and Bale, 2014). In the most severe forms, such as harlequin ichthyosis, the disease is often fatal during the early postnatal period, whereas at the other end of the continuum of the spectrum, the disease may present with a relatively mild scaling and variable degree of palmoplantar keratoderma. There is considerable genetic heterogeneity in ARCI, and as many as nine different genes are known to harbor biallelic mutations; these include TGM1, ALOXE3, ALOX12B, NIPAL4, ABCA12, CYP4F22, PNPLA1, LIPN, and CERS3. Previous reports have suggested that mutations in TGM1 account for 30e65% of patients with ARCI, whereas mutations in LIPN, PNPLA1, and CERS3 have been reported only in a few consanguineous families (Richard and Bale, 2014). With the advent of next generation sequencing (NGS), there has been tremendous progress in facilitating the mutation detection in various heritable skin disorders, including ichthyosis (South et al., 2015; Takeichi et al., 2013). In fact, at least 38 different genes have now been suggested to be associated with the ichthyotic phenotypes, either as the primary mutated genes or modifying the phenotypic presentation. To elucidate the genetic basis of ichthyosis in Iran, a country of approximately 80 million people with high prevalence of customary consanguineous marriages, we developed a gene-targeted NGS array consisting of 38 genes reported in association with ichthyosis phenotypes. Identification of specific mutations in a large number of families has allowed us to examine phenotype/genotype correlations with respect to both intra- and interfamilial heterogeneity, in part because of extensive consanguinity in these families. In this study, we identified six distinct and, to our knowledge, previously unreported mutations in the PNPLA1 gene in nine families

Multigene Next-Generation Sequencing Panel Identifies Pathogenic Variants in Patients with Unknown Subtype of Epidermolysis Bullosa: Subclassification with Prognostic Implications

Purpose: Epidermolysis bullosa (EB), the prototype of heritable blistering diseases, is caused by mutations in as many as 19 distinct genes. In this study, we evaluated the molecular basis of EB in 93 families, many of them of unknown subtype. Methods: A next generation sequencing panel covering 21 EB-related genes was developed, and mutation profiles, together with clinical features, were used to classify the patients into distinct clinical categories. Results: A total of 72 pathogenic or likely pathogenic variants in 68 families were identified in 11 of the EB-associated candidate genes, most of them (75%) being homozygous consistent with considerable consanguinity in the cohort. Approximately half of the mutations (48.6%) were previously unreported. Refined analysis of the types of mutations with addition of variants of unknown significance suspected of causing clinically consistent disease in several patients allowed subclassification of patients into different subcategories of EB in 76 of 91 families, with prognostic implications. A genetically challenging case with homozygous loss-of-function pathogenic mutations in two different EB-associated genes resulting in different subtypes was identified. In addition, secondary findings included identification of known pathogenic variants in DSP associated with arrhythmogenic right ventricular cardiomyopathy. Conclusion: Utilization of next generation sequencing panel of EB-associated genes allowed diagnostic subclassification in the neonates particularly in families of unknown subtype, with prognostication of the overall long-term outcome of the disease

Recessive mutation in tetraspanin CD151 causes Kindler syndrome-like epidermolysis bullosa with multi-systemic manifestations including nephropathy

Epidermolysis bullosa (EB) is caused by mutations in as many as 19 distinct genes. We have developed a next-generation sequencing (NGS) panel targeting genes known to be mutated in skin fragility disorders, including tetraspanin CD151 expressed in keratinocytes at the dermal-epidermal junction. The NGS panel was applied to a cohort of 92 consanguineous families of unknown subtype of EB. In one family, a homozygous donor splice site mutation in CD151 (NM_139029; c.351 + 2T > C) at the exon 5/intron 5 border was identified, and RT-PCR and whole transcriptome analysis by RNA-seq confirmed deletion of the entire exon 5 encoding 25 amino acids. Immunofluorescence of proband's skin and Western blot of skin proteins with a monoclonal antibody revealed complete absence of CD151. Transmission electron microscopy showed intracellular disruption and cell-cell dysadhesion of keratinocytes in the lower epidermis. Clinical examination of the 33-year old proband, initially diagnosed as Kindler syndrome, revealed widespread blistering, particularly on pretibial areas, poikiloderma, nail dystrophy, loss of teeth, early onset alopecia, and esophageal webbing and strictures. The patient also had history of nephropathy with proteinuria. Collectively, the results suggest that biallelic loss-of-function mutations in CD151 underlie an autosomal recessive mechano-bullous disease with systemic features. Thus, CD151 should be considered as the 20th causative, EB-associated gene

Hypotrichosis with juvenile macular dystrophy: Combination of whole-genome sequencing and genome-wide homozygosity mapping identifies a large deletion in CDH3 initially undetected by whole-exome sequencing-A lesson from next-generation sequencing.

BACKGROUND: Hypotrichosis with juvenile macular dystrophy (HJMD) is an autosomal recessive disorder characterized by abnormal growth of scalp hair and juvenile macular degeneration leading to blindness. We have explored the genetic basis of HJMD in a large consanguineous family with 12 affected patients, 1-76 years of age, with characteristic phenotypes. METHODS: We first applied genome-wide homozygosity mapping to 10 affected individuals for linkage analysis to identify the genomic region of the defective gene. All affected individuals shared a 7.2 Mb region of homozygosity on chromosome 16q21-22.3, which harbored 298 genes, including CDH3, previously associated with HJMD. However, whole-exome sequencing (WES) failed to identify the causative mutation in CDH3. RESULTS: Further investigation revealed a missense variant in a gene closely linked to CDH3 (1.4 Mb distance: FHOD1: c.1306A\u3eG, p.Arg436Gly). This variant was homozygous in all affected individuals and heterozygous in 18 out of 19 obligate carriers. While this variant was found by bioinformatics predictions to be likely pathogenic, a knock-in mouse for this variant, made by the CRISPR/Cas, showed no disease phenotype. However, using whole-genome sequencing (WGS), we were able to identify a novel Alu recombination-mediated deletion in CDH3:c.del161-811_246 + 1,044. CONCLUSION: WGS was able to identify a deep intronic deletion mutation, not detected by WES

High-Throughput Label-Free Isolation of Heterogeneous Circulating Tumor Cells and CTC Clusters from Non-Small-Cell Lung Cancer Patients.

(1) Background: Circulating tumor cell (CTC) clusters are emerging as clinically significant harbingers of metastases in solid organ cancers. Prior to engaging these CTC clusters in animal models of metastases, it is imperative for technology to identify them with high sensitivity. These clusters often present heterogeneous surface markers and current methods for isolation of clusters may fall short. (2) Methods: We applied an inertial microfluidic Labyrinth device for high-throughput, biomarker-independent, size-based isolation of CTCs/CTC clusters from patients with metastatic non-small-cell lung cancer (NSCLC). (3) Results: Using Labyrinth, CTCs (PanCK+/DAPI+/CD45-) were isolated from patients (n = 25). Heterogeneous CTC populations, including CTCs expressing epithelial (EpCAM), mesenchymal (Vimentin) or both markers were detected. CTCs were isolated from 100% of patients (417 +/- 1023 CTCs/mL). EpCAM- CTCs were significantly greater than EpCAM+ CTCs. Cell clusters of \u3e/=2 CTCs were observed in 96% of patients-of which, 75% were EpCAM-. CTCs revealed identical genetic aberrations as the primary tumor for RET, ROS1, and ALK genes using fluorescence in situ hybridization (FISH) analysis. (4) Conclusions: The Labyrinth device recovered heterogeneous CTCs in 100% and CTC clusters in 96% of patients with metastatic NSCLC. The majority of recovered CTCs/clusters were EpCAM-, suggesting that these would have been missed using traditional antibody-based capture methods