Copy Number Variation Analysis Research Articles

Genetic Association Analysis of Copy Number Variations for Meat Quality in Beef Cattle.

Meat quality is an economically important trait for global food production. Copy number variations (CNVs) have been previously implicated in elucidating the genetic basis of complex traits. In this article, we detected a total of 112,198 CNVs and 10,102 CNV regions (CNVRs) based on the Bovine HD SNP array. Next, we performed a CNV-based genome-wide association analysis (GWAS) of six meat quality traits and identified 12 significant CNV segments corresponding to eight candidate genes, including PCDH15, CSMD3, etc. Using region-based association analysis, we further identified six CNV segments relevant to meat quality in beef cattle. Among these, TRIM77 and TRIM64 within CNVR4 on BTA29 were detected as candidate genes for backfat thickness (BFT). Notably, we identified a 34 kb duplication for meat color (MC) which was supported by read-depth signals, and this duplication was embedded within the keratin gene family including KRT4, KRT78, and KRT79. Our findings will help to dissect the genetic architecture of meat quality traits from the aspects of CNVs, and subsequently improve the selection process in breeding programs.

Cat eye syndrome caused by 22q11.1q11.21 duplication: case report in a Chinese family

Purpose This paper presents a report on two uncommon instances of cat eye syndrome in a Chinese family.Case presentationThe proband, a 23-year-old female, exhibited a diminutive cornea and complete blindness in her right eye, and the uncorrected distance visual acuity of her left eye was 0.7 LogMAR. Peripheral blood chromosome karyotyping reveal a karyotype of 47, XX, + mar. Subsequent analysis of chromosome copy number variation unveiled a 1.5 Mb duplication in the 22q11.1q11.21 region of the proband. The proband's mother,aged 49, displayed small eyes, wide-set eyes, downward slanting eyelids, and congenital heart disease. Chromosome copy number variation analysis also showed a 1.55 Mb duplication in the 22q11.1q11.21 region of chromosome 22 in the proband's mother. Ultimately, both members of this family were diagnosed with cat eye syndrome.Conclusion Cat eye syndrome is a rare genetic disorder that greatly affects patients' lives and requires personalized treatment. This study provides new evidence for a better understanding of the diagnosis of cat eye syndrome and emphasizes the importance of genetic counseling and supervision.

Mesonephric-like adenocarcinoma harbours characteristic copy number variations and a distinct DNA methylation signature closely related to mesonephric adenocarcinoma of the cervix.

Mesonephric-like adenocarcinoma (MLA) of the female genital tract is an uncommon histotype that can arise in both the endometrium and the ovary. The exact cell of origin and histogenesis currently remain unknown. Here, we investigated whole genome DNA methylation patterns and copy number variations (CNVs) in a series of MLAs in the context of a large cohort of various gynaecological carcinoma types. CNV analysis of 19 MLAs uncovered gains of chromosomes 1q (18/19, 95%), 10 (15/19, 79%), 12 (14/19, 74%), and 2 (10/19, 53%), as well as loss of chromosome 1p (7/19, 37%). Gains of chromosomes 1q, 10, and 12 were also identified in the majority of mesonephric adenocarcinomas of the uterine cervix (MAs) as well as subsets of endometrioid carcinomas (ECs) and low-grade serous carcinomas of the ovary (LGSCs) but only in a minority of serous carcinomas of the uterine corpus (USCs), clear cell carcinomas (CCCs), and tubo-ovarian high-grade serous carcinomas (HGSCs). While losses of chromosome 1p together with gains of chromosome 1q were also identified in both MA and LGSC, gains of chromosome 2 were almost exclusively identified in MLA and MA. Unsupervised hierarchical clustering and t-SNE analysis of DNA methylation data (Illumina EPIC array) identified a co-clustering for MLAs and MAs, which was distinct from clusters of ECs, USCs, CCCs, LGSCs, and HGSCs. Group-wise comparisons confirmed a close epigenetic relationship between MLA and MA. These findings, in conjunction with the established histological and immunophenotypical overlap, suggest bona fide mesonephric differentiation, and support a more precise terminology of mesonephric-type adenocarcinoma instead of MLA in these tumours. © 2023 The Authors. The Journal of Pathology published by John Wiley & Sons Ltd on behalf of The Pathological Society of Great Britain and Ireland.

Analysis of copy number variations and possible candidate genes in spontaneous abortion by copy number variation sequencing.

Embryonic chromosomal abnormalities represent a major causative factor in early pregnancy loss, highlighting the importance of understanding their role in spontaneous abortion. This study investigates the potential correlation between chromosomal abnormalities and spontaneous abortion using copy number variation sequencing (CNV-seq), a Next-Generation Sequencing (NGS) technology. We analyzed Copy Number Variations (CNVs) in 395 aborted fetal specimens from spontaneous abortion patients by CNV-seq. And collected correlated data, including maternal age, gestational week, and Body Mass Index (BMI), and analyzed their relationship with the CNVs. Out of the 395 cases, 67.09% of the fetuses had chromosomal abnormalities, including numerical abnormalities, structural abnormalities, and mosaicisms. Maternal age was found to be an important risk factor for fetal chromosomal abnormalities, with the proportion of autosomal trisomy in abnormal karyotypes increasing with maternal age, while polyploidy decreased. The proportion of abnormal karyotypes with mosaic decreased as gestational age increased, while the frequency of polyploidy and sex chromosome monosomy increased. Gene enrichment analysis identified potential miscarriage candidate genes and functions, as well as pathogenic genes and pathways associated with unexplained miscarriage among women aged below or over 35 years old. Based on our study, it can be inferred that there is an association between BMI values and the risk of recurrent miscarriage caused by chromosomal abnormalities. Overall, these findings provide important insights into the understanding of spontaneous abortion and have implications for the development of personalized interventions for patients with abnormal karyotypes.

Genome-Wide Detection of Copy Number Variations Associated with Miniature Features in Horses.

Copy number variations (CNVs) are crucial structural genomic variants affecting complex traits in humans and livestock animals. The current study was designed to conduct a comprehensive comparative copy number variation analysis among three breeds, Debao (DB), Baise (BS), and Warmblood (WB), with a specific focus on identifying genomic regions associated with miniature features in horses. Using whole-genome next-generation resequencing data, we identified 18,974 CNVs across 31 autosomes. Among the breeds, we found 4279 breed-specific CNV regions (CNVRs). Baise, Debao, and Warmblood displayed 2978, 986, and 895 distinct CNVRs, respectively, with 202 CNVRs shared across all three breeds. After removing duplicates, we obtained 1545 CNVRs from 26 horse genomes. Functional annotation reveals enrichment in biological functions, including antigen processing, cell metabolism, olfactory conduction, and nervous system development. Debao horses have 970 genes overlapping with CNVRs, possibly causing their small size and mountainous adaptations. We also found that the genes GHR, SOX9, and SOX11 may be responsible for the miniature features of the Debao horse by analyzing their overlapping CNVRs. Overall, this study offers valuable insights into the widespread presence of CNVs in the horse genome. The findings contribute to mapping horse CNVs and advance research on unique miniature traits observed in the Debao horse.

STS and PUDP Deletion Identified by Targeted Panel Sequencing with CNV Analysis in X-Linked Ichthyosis: A Case Report and Literature Review.

X-linked recessive ichthyosis (XLI) is clinically characterized by dark brown, widespread dryness with polygonal scales. We describe the identification of STS and PUDP deletions using targeted panel sequencing combined with copy-number variation (CNV) analysis in XLI. A 9-month-old infant was admitted for genetic counseling. Since the second day after birth, the infant's skin tended to be dry and polygonal scales had accumulated over the abdomen and upper extremities. The infant's maternal uncle and brother (who had also exhibited similar skin symptoms from birth) presented with polygonal scales on their trunks. CNV analysis revealed a hemizygous deletion spanning 719.3 Kb on chromosome Xp22 (chrX:7,108,996-7,828,312), which included a segment of the STS gene and exhibited a Z ratio of -2 in the proband. Multiplex ligation-dependent probe amplification (MLPA) confirmed this interstitial Xp22.31 deletion. Our report underscores the importance of implementing CNV screening techniques, including sequencing data analysis and gene dosage assays such as MLPA, to detect substantial deletions that encompass the STS gene region of Xq22 in individuals suspected of having XLI.

Genomic Variations Explorer (GenVarX): a toolset for annotating promoter and CNV regions using genotypic and phenotypic differences

The rapid growth of sequencing technology and its increasing popularity in biology-related research over the years has made whole genome re-sequencing (WGRS) data become widely available. A large amount of WGRS data can unlock the knowledge gap between genomics and phenomics through gaining an understanding of the genomic variations that can lead to phenotype changes. These genomic variations are usually comprised of allele and structural changes in DNA, and these changes can affect the regulatory mechanisms causing changes in gene expression and altering the phenotypes of organisms. In this research work, we created the GenVarX toolset, that is backed by transcription factor binding sequence data in promoter regions, the copy number variations data, SNPs and Indels data, and phenotypes data which can potentially provide insights about phenotypic differences and solve compelling questions in plant research. Analytics-wise, we have developed strategies to better utilize the WGRS data and mine the data using efficient data processing scripts, libraries, tools, and frameworks to create the interactive and visualization-enhanced GenVarX toolset that encompasses both promoter regions and copy number variation analysis components. The main capabilities of the GenVarX toolset are to provide easy-to-use interfaces for users to perform queries, visualize data, and interact with the data. Based on different input windows on the user interface, users can provide inputs corresponding to each field and submit the information as a query. The data returned on the results page is usually displayed in a tabular fashion. In addition, interactive figures are also included in the toolset to facilitate the visualization of statistical results or tool outputs. Currently, the GenVarX toolset supports soybean, rice, and Arabidopsis. The researchers can access the soybean GenVarX toolset from SoyKB via https://soykb.org/SoybeanGenVarX/, rice GenVarX toolset, and Arabidopsis GenVarX toolset from KBCommons web portal with links https://kbcommons.org/system/tools/GenVarX/Osativa and https://kbcommons.org/system/tools/GenVarX/Athaliana, respectively.

Low-pass whole genome sequencing is a reliable and cost-effective approach for copy number variant analysis in the clinical setting.

Next generation sequencing technology has greatly reduced the cost and time required for sequencing a genome. An approach that is rapidly being adopted as an alternative method for CNV analysis is the low-pass whole genome sequencing (LP-WGS). Here, we evaluated the performance of LP-WGSto detect copy number variants (CNVs) in clinical cytogenetics. DNA samples with known CNVs detected by chromosomal microarray analyses (CMA) were selected for comparison and used as positive controls; our panel included 44 DNA samples (12 prenatal and 32 postnatal), comprising a total of 55 chromosome imbalances. The selected cases were chosen to provide a wide range of clinically relevant CNVs, the vast majority being associated with intellectual disability or recognizable syndromes. The chromosome imbalances ranged in size from 75kb to 90.3Mb, including aneuploidies and two cases of mosaicism. All CNVs were successfully detected by LP-WGS, showing a high level of consistency and robust performance of the sequencing method. Notably, the size of chromosome imbalances detected by CMA and LP-WGS were compatible between the two different platforms, which indicates that the resolution and sensitivity of the LP-WGS approach are at least similar to those provided by CMA. Our data show the potential use of LP-WGS to detect CNVs in clinical diagnosis and confirm the method as an alternative for chromosome imbalances detection. The diagnostic effectiveness and feasibility of LP-WGS, in this technical validation study, were evidenced by a clinically representative dataset of CNVs that allowed a systematic assessment of the detection power and the accuracy of the sequencing approach. Further, since the software used in this study is commercially available, the method can easily be tested and implemented in a routine diagnostic setting.

Significant decrease of maternal mitochondria carryover using optimized spindle-chromosomal complex transfer.

Mutations in mitochondrial DNA (mtDNA) contribute to a variety of serious multi-organ human diseases, which are strictly inherited from the maternal germline. However, there is currently no curative treatment. Attention has been focused on preventing the transmission of mitochondrial diseases through mitochondrial replacement (MR) therapy, but levels of mutant mtDNA can often unexpectedly undergo significant changes known as mitochondrial genetic drift. Here, we proposed a novel strategy to perform spindle-chromosomal complex transfer (SCCT) with maximal residue removal (MRR) in metaphase II (MII) oocytes, thus hopefully eliminated the transmission of mtDNA diseases. With the MRR procedure, we initially investigated the proportions of mtDNA copy numbers in isolated karyoplasts to those of individual oocytes. Spindle-chromosomal morphology and copy number variation (CNV) analysis also confirmed the safety of this method. Then, we reconstructed oocytes by MRR-SCCT, which well developed to blastocysts with minimal mtDNA residue and normal chromosomal copy numbers. Meanwhile, we optimized the manipulation order between intracytoplasmic sperm injection (ICSI) and SCC transfer and concluded that ICSI-then-transfer was conducive to avoid premature activation of reconstructed oocytes in favor of normal fertilization. Offspring of mice generated by embryos transplantation in vivo and embryonic stem cells derivation further presented evidences for competitive development competence and stable mtDNA carryover without genetic drift. Importantly, we also successfully accomplished SCCT in human MII oocytes resulting in tiny mtDNA residue and excellent embryo development through MRR manipulation. Taken together, our preclinical mouse and human models of the MRR-SCCT strategy not only demonstrated efficient residue removal but also high compatibility with normal embryo development, thus could potentially be served as a feasible clinical treatment to prevent the transmission of inherited mtDNA diseases.

OR21-02 Diagnostic Yield Of A Multigene Approach In Children With Short Stature Of Unknown Cause

Abstract Disclosure: N.L. Menezes De Andrade: None. M.F. Funari: None. A. Malaquias: Speaker; Self; Novo Nordisk. N.C. Dantas: None. R. Rezende: None. L.D. Cellin: None. T.K. Homma: None. A.M. Lerario: None. I.J. Arnhold: None. G.A. Vasques: Employee; Self; Eli Lilly & Company. A.A. Jorge: Advisory Board Member; Self; Novo Nordisk. Consulting Fee; Self; Novo Nordisk. Research Investigator; Self; Novo Nordisk. Speaker; Self; Pfizer Global R&D. Introduction: The genetic factors are frequently implicated in the growth impairment of short stature (SS) children, whether they are syndromic or apparently healthy. However, the investigation recommended by the consensus fails to establish the etiology in most cases. The difficulty to search for candidate genes increases the importance of using a hypothesis-free approach as whole exome sequencing (WES) for these children. Objective: To determine the diagnostic yield of a multigene gene analysis in a cohort of children with SS of unknown cause. Patients and methods: We enrolled children with syndromic (n= 143) and isolated SS (n=380) of unknown cause after traditional investigation. The genetic analysis was applied to all patients [WES =306, panel = 217, including copy number variants (CNVs) analysis]. A chromosomal microarray analysis was also used for 79 syndromic SS. Results: A genetic defect was identified in 74 and 70 children with syndromic and isolated SS, respectively. Variants associated with syndromic SS were widely distributed in several rare conditions, affecting 63 different genes: COL2A1 in 3 patients; ANKRD11; BCL11B; LTBP3; CHD7; PTPN11; PUF60; SRCAP; STAT5B, GNAS in two patients each; SCN1A; SETD5; PTHLH; GINS1; ZMYM2; TERT; G6PC3; KIF11; SCUBE3; TBC1D32; PDHA1; RPS6KA3; SMARCA4; DNM2; KMT2A ; IHH; BRCA1; NF1; INPP5K; KRAS; FGFR2; DPH1; RPL5; KDM5C; DYRK1A; MVK; FBN1; AFF4; ACTB; POC1A; IGF1R; CREBBP; POLD1; POLR3B; PCNA; CUL7; RUNX2; EXT1; GH1; FGD1; BLM; CHRNG in one patient each. In children with isolated SS, 5 children had more than one causative finding. There were 39 defects associated with the growth plate [SHOX (n=13); NPR2 (n=7); IHH (n=10); ACAN (n=5); FGFR3 (n=3); COL2A1 (n=1)], 6 involved in the GH-IGF-1 axis [GHSR (n=3); IGF1R (n=2); GH1 (n=1)], 9 related to the RAS/MAPK pathway [PTPN11 (n=4); NF1 (n=3); BRAF (n=1); CBL (n=1)]; and 17 other genes [LTBP3 (n=2); THRA (n=2); TYMP; PIK3CA; CDKN1C; ANKRD11; HMGA2; SRCAP; KMT2C; GDF5; OBSL1; MC4R; FBN1; PRKG2; ERF (one patient each)]. Additionally, a pathogenic CNVs were identified in 11 and 3 patients with syndromic and isolated SS, respectively. The diagnostic rate was 51.7% for patients with syndromic features vs 18,4% in patients with isolated SS (p < 0.001). Patients with or without a genetic diagnosis cannot be clinically differentiated into either group. Conclusion: The diagnostic yield of a multigene sequencing approach was three time higher in patients with syndromic SS than those with isolated SS. Nevertheless, the identification of a monogenic condition has a clear impact on patients’ follow-up and treatment for both groups. Maximum of 2,500 characters, including punctuation (not spaces). Presentation: Saturday, June 17, 2023

Specific genetic aberrations of parathyroid in Chinese patients with tertiary hyperparathyroidism using whole-exome sequencing.

Tertiary hyperparathyroidism (THPT) is a peculiar subtype of hyperparathyroidism that usually develops from chronic kidney disease (CKD) and persists even after kidney transplantation. Unlike its precursor, secondary hyperparathyroidism (SHPT), THPT is characterized by uncontrolled high levels of calcium in the blood, which suggests the monoclonal or oligoclonal proliferation of parathyroid cells. However, the molecular abnormalities leading to THPT have not yet been fully understood. In this study, we analyzed DNA samples from hyperplastic parathyroid and corresponding blood cells of 11 patients with THPT using whole-exome sequencing (WES). We identified somatic single nucleotide variants (SNV) and insertions or deletions variants (INDEL) and performed driver mutation analysis, KEGG pathway, and GO functional enrichment analysis. To confirm the impact of selected driver mutated genes, we also tested their expression level in these samples using qRT-PCR. Following quality control and mutation filtering, we identified 17,401 mutations, comprising 6690 missense variants, 3078 frameshift variants, 2005 stop-gained variants, and 1630 synonymous variants. Copy number variants (CNV) analysis showed that chromosome 22 copy number deletion was frequently observed in 6 samples. Driver mutation analysis identified 179 statistically significant mutated genes, including recurrent missense mutations on TBX20, ATAD5, ZNF669, and NOX3 genes in 3 different patients. KEGG pathway analysis revealed two enriched pathways: non-homologous end-joining and cell cycle, with a sole gene, PRKDC, involved. GO analysis demonstrated significant enrichment of various cellular components and cytobiological processes associated with four genes, including GO items of positive regulation of developmental growth, protein ubiquitination, and positive regulation of the apoptotic process. Compared to blood samples, THPT samples exhibited lower expression levels of PRKDC, TBX20, ATAD5, and NOX3 genes. THPT samples with exon mutations had relatively lower expression levels of PRKDC, TBX20, and NOX3 genes compared to those without mutations, although the difference was not statistically significant. This study provides a comprehensive landscape of the genetic characteristics of hyperplastic parathyroids in THPT, highlighting the involvement of multiple genes and pathways in the development and progression of this disease. The dominant mutations identified in our study depicted new insights into the pathogenesis and molecular characteristics of THPT.

A comprehensive pan-cancer analysis identifies a novel glycolysis score and its hub genes as prognostic and immunological biomarkers.

Glycolysis plays significant roles in tumor progression and immune response. However, the exact role of glycolysis in prognosis and immune regulation has not been explored in all cancer types. This study first calculated a novel glycolysis score and screened out 12 glycolytic hub genes, and comprehensively analyzed molecular expression, clinical implications, and immune features of glycolysis among pan-cancer. The glycolysis score was derived by the single sample gene set enrichment analysis (ssGSEA) algorithm. The correlations of glycolysis with clinical parameters were analyzed using "limma" package. Downstream pathways of glycolysis were identified by Gene Set Enrichment Analysis (GSEA). The immune cell infiltration was explored and validated by three databases. The association between glycolysis and some immunotherapy biomarkers was explored by Pearson correlation analysis. Single-nucleotide variation (SNV), copy number variation (CNV), DNA methylation, and drug sensitivity analyses of 12 glycolytic hub genes were investigated. IMvigor210 and GSE91061 immunotherapeutic cohorts were retrieved to assess the ability of glycolysis score to predict immunotherapy efficacy. The expression of glycolysis key genes was detected in normal and endometrial cancer cell lines. We found that glycolysis score was generally higher in tumor tissues compared to normal tissues and a high glycolysis score predominated as a risk prognostic factor. A high glycolysis score was associated with decreased immunostimulatory natural killer (NK) cells and CD8+ T cells infiltration, well increased immunosuppressive M2-tumor-associated macrophages (M2-TAM) cells infiltration. Tumor mutational burden (TMB), microsatellite instability (MSI), and immune checkpoints (ICPs) all closely interacted with glycolysis score and the frequency of gene mutation was confirmed to be higher in colon adenocarcinoma (COAD) patients with higher glycolysis score. The SNV, CNV, and DNA methylation of 12 glycolysis key genes occurred at different frequencies and showed different impacts on survival outcomes. The predictive and prognostic value of glycolysis score for immunotherapy outcomes was validated in two immunotherapy cohorts. The expression levels of key genes differ in normal endometrial and three endometrial cancer cell lines. This work indicated that glycolysis score and 12 glycolytic hub genes were correlated with an immunosuppressive microenvironment. They could be served as promising biomarkers aiding diagnosis, predicting prognosis and immunotherapy response for some tumor patients.

Identification of Cell Type-Specific Effects of DNMT3A Mutations on Relapse in Acute Myeloid Leukemia.

Acute myeloid leukemia (AML) is a heterogeneous disease caused by distinctive mutations in individual patients; therefore, each patient may display different cell-type compositions. Although most patients with AML achieve complete remission (CR) through intensive chemotherapy, the likelihood of relapse remains high. Several studies have attempted to characterize the genetic and cellular heterogeneity of AML; however, our understanding of the cellular heterogeneity of AML remains limited. In this study, we performed single-cell RNA sequencing (scRNAseq) of bone marrow-derived mononuclear cells obtained from same patients at different AML stages (diagnosis, CR, and relapse). We found that hematopoietic stem cells (HSCs) at diagnosis were abnormal compared to normal HSCs. By improving the detection of the DNMT3A R882 mutation with targeted scRNAseq, we identified that DNMT3A-mutant cells that mainly remained were granulocyte-monocyte progenitors (GMPs) or lymphoid-primed multipotential progenitors (LMPPs) from CR to relapse and that DNMT3A-mutant cells have gene signatures related to AML and leukemic cells. Copy number variation analysis at the single-cell level indicated that the cell type that possesses DNMT3A mutations is an important factor in AML relapse and that GMP and LMPP cells can affect relapse in patients with AML. This study advances our understanding of the role of DNMT3A in AML relapse and our approach can be applied to predict treatment outcomes.

A novel microdeletion of 517 kb downstream of the PAX6 gene in a Chinese family with congenital aniridia

BackgroundTo identify the disease-causing gene in a Chinese family affected with congenital aniridia.MethodsPatients underwent systematic ophthalmic examinations such as anterior segment photography, fundus photography, optical coherence tomography, and fundus fluorescein angiography. The proband was screened for pathogenic variants by whole exome sequencing (WES) and copy number variant (CNV) analysis. Real-time quantitative PCR (RT-qPCR) was applied to confirm the CNV results. Breakpoints were identified by long-range PCR followed by Sanger sequencing.ResultsAll seven members of this Chinese family, including four patients and three normal individuals, were recruited for this study. All patients showed bilateral congenital aniridia with nystagmus, except the son of the proband, who presented with bilateral partial coloboma of the iris. A novel heterozygous deletion (chr11:31,139,019–31,655,997) containing the 3’ regulatory enhancers of the PAX6 gene was detected in this family. We also reviewed the reported microdeletions downstream of PAX6 in patients with aniridia.ConclusionsWe identified a novel microdeletion, 517 kb in size located about 133 kb downstream of the PAX6 gene, responsible for congenital aniridia in this Chinese family, which expands the spectrum of aniridia-associated mutations in PAX6.

A novel embryo biopsy morphological analysis and genetic integrality criterion system significantly improves the outcome of preimplantation genetic testing.

Whileefforts have been made to establish blastocyst grading systems in the past decades, little research has examined the quality of biopsy specimens. This study is the first to correlate the morphology of biopsied trophectoderm (TE) cells to their quality and subsequent genetic testing results of preimplantation genetic testing (PGT), through an innovative Morphological Analysis and Genetic Integrality Criterion (MAGIC) system. Biopsied TE cells were first evaluated according to the MAGIC procedure, followed by whole-genome amplification (WGA) and library construction, and then sequenced using the Illumina X Ten Platform. Copy number variation (CNV) and allele drop-out (ADO) rates as well as test failure rates were compared and analyzed. Our data explores the relationship between TE cell morphology and its quality and final genetic testing outcome, which is established based on the MAGIC system. MAGIC guarantees that only high- or good-quality TE cells are used for genetic testing to generate excellent data uniformity and lower ADO rates. Low-quality cells containing biopsied TE cell mass are responsible for the "background noise" of CNV analysis. The MAGIC application has effectively decreased the false-positive mosaicism, hence to ensure the stability and veracity of detection results, to avoid misdiagnoses, and to improve accuracy, as well as to avoid re-biopsy procedures. The study also contributes to understand how the IVF laboratory and the molecular biology laboratory depend on each other to achieve good-quality PGT results, which are clinically relevant for the patients.

Different Genetic Signatures of Small-Cell Lung Cancer Characterize Anti-GABAB R and Anti-Hu Paraneoplastic Neurological Syndromes.

Small-cell lung cancer (SCLC) is the malignancy most frequently associated with paraneoplastic neurological syndromes (PNS) and can trigger different antibody responses against intracellular (Hu) or neuronal surface (GABAB R) antigens. Our aim was to clarify whether the genomic and transcriptomic features of SCLC are different in patients with anti-GABAB R or anti-Hu PNS compared with SCLC without PNS. A total of 76 SCLC tumor samples were collected: 34 anti-Hu, 14 anti-GABAB R, and 28 SCLC without PNS. The study consisted of 4 steps: (1) pathological confirmation; (2) next generation sequencing using a panel of 98 genes, including those encoding the autoantibodies targets ELAVL1-4, GABBR1-2, and KCTD16; (3) genome-wide copy number variation (CNV); and (4) whole-transcriptome RNA sequencing. CNV analysis revealed that patients with anti-GABAB R PNS commonly have a gain in chromosome 5q, which contains KCTD16, whereas anti-Hu and control patients often harbor a loss. No significantly different number of mutations regarding any onconeural genes was observed. Conversely, the transcriptomic profile of SCLC was different, and the differentially expressed genes allowed effective clustering of the samples into 3 groups, reflecting the antibody-based classification, with an overexpression of KCTD16 specific to anti-GABAB R PNS. Pathway analysis revealed that tumors of patients with anti-GABAB R encephalitis were enriched in B-cell signatures, as opposed to those of patients with anti-Hu, in which T-cell- and interferon-γ-related signatures were overexpressed. SCLC genetic and transcriptomic features differentiate anti-GABAB R, anti-Hu, and non-PNS tumors. The role of KCTD16 appears to be pivotal in the tumor immune tolerance breakdown of anti-GABAB R PNS. ANN NEUROL 2023;94:1102-1115.

Genetic diagnostic yield in an 11-year cohort of craniosynostosis patients

Craniosynostosis may present in isolation, ‘non-syndromic’, or with additional congenital anomalies/neurodevelopmental disorders, ‘syndromic’. Clinical focus shifted from confirming classical syndromic cases to offering genetic testing to all craniosynostosis patients. This retrospective study assesses diagnostic yield of molecular testing by investigating prevalences of chromosomal and monogenic (likely) pathogenic variants in an 11-year cohort of 1020 craniosynostosis patients. 502 children underwent genetic testing. Pathogenic variants were identified in 174 patients (35%). Diagnostic yield was significantly higher in syndromic craniosynostosis (62%) than in non-syndromic craniosynostosis (6%). Before whole exome sequencing (WES) emerged, single-gene testing was performed using Sanger sequencing or multiplex ligation-dependent probe amplification (MLPA). Diagnostic yield was 11% and was highest for EFNB1, FGFR2, FGFR3, and IL11RA. Diagnostic yield for copy number variant analysis using microarray was 8%. From 2015 onwards, the WES craniosynostosis panel was implemented, with a yield of 10%. In unsolved, mainly syndromic, cases suspected of a genetic cause, additional WES panels (multiple congenital anomalies (MCA)/intellectual disability (ID)) or open exome analysis were performed with an 18% diagnostic yield. To conclude, microarray and the WES craniosynostosis panel are key to identifying pathogenic variants. in craniosynostosis patients. Given the advances in genetic diagnostics, we should look beyond the scope of the WES craniosynostosis panel and consider extensive genetic diagnostics (e.g. open exome sequencing, whole genome sequencing, RNA sequencing and episignature analysis) if no diagnosis is obtained through microarray and/or WES craniosynostosis panel. If parents are uncomfortable with more extensive diagnostics, MCA or ID panels may be considered.

Identification of the first homozygous intragenic deletion in the YY1AP1 gene in a consanguineous family: New insights into the phenotypic variability associated with Grange syndrome.

Grange syndrome (GRNG-MIM#135580) is a rare recessive disorder associating variable features including diffuse vascular stenosis, brachysyndactyly, osteopenia with increased bone fragility, cardiac malformations, and variable developmental delay. Since its first description in 1998, only 15 individuals from 10 families have been reported, carrying homozygous or compound heterozygous frameshift or nonsense variants in YY1AP1. In a patient with cutaneous and bone syndactyly and a hemorrhagic stroke at the age of 16 months, consistent with a clinical diagnosis of GRNG, we performed exome sequencing after negative array-CGH and congenital limb malformation panel results. Copy number variant analysis from exome data identified a homozygous intragenic out-of-frame deletion of 1.84 kb encompassing exons seven and eight of YY1AP1, confirming a molecular diagnosis of GRNG. Genetic counseling led to the identification of additional family members compatible with GRNG. Here, we provide new insights into the phenotypic variability associated with GRNG and highlight the utility of the detection of small copy number variants to identify the molecular causes of heterogeneous malformative genetic disorders.

Comprehensive Genome and Transcriptome Analysis Identifies SLCO3A1 Associated with Aggressive Behavior in Pigs.

Copy number variation (CNV) represents a significant reservoir of genetic diversity within the genome and exhibits a strong association with economically valuable traits in livestock. The manifestation of aggressive behavior in pigs has detrimental effects on production efficiency, immune competency, and meat quality. Nevertheless, the impact of CNV on the aggressive behavior of pigs remains elusive. In this investigation, we employed an integrated analysis of genome and transcriptome data to investigate the interplay between CNV, gene expression changes, and indicators of aggressive behavior in weaned pigs. Specifically, a subset of pigs comprising the most aggressive pigs (MAP, n = 12) and the least aggressive pigs (LAP, n = 11) was purposefully selected from a herd of 500 weaned pigs following a mixing procedure based on their composite aggressive score (CAS). Subsequently, we thoroughly analyzed copy number variation regions (CNVRs) across the entire genome using next-generation sequencing techniques, ultimately revealing the presence of 6869 CNVRs. Using genome-wide association study (GWAS) analysis and evaluating variance-stabilizing transformation (VST) values, we successfully identified distinct CNVRs that distinguished the MAP and LAP counterparts. Among the prioritized CNVRs, CNVR-4962 (designated as the top-ranked p-value and VST value, No. 1) was located within the Solute Carrier Organic Anion Transporter Family Member 3A1 (SLCO3A1) gene. The results of our analyses indicated a significantly higher (p < 0.05) copy number of SLCO3A1 in the MAP compared to the LAP. Furthermore, this increased copy number exhibited a positive correlation with the CAS of the pigs (p < 0.05). Furthermore, we integrated genomic data with transcriptomic data from the temporal lobe to facilitate the examination of expression quantitative trait loci (eQTL). Importantly, these observations were consistent with the mRNA expression pattern of SLCO3A1 in the temporal lobe of both MAP and LAP (p < 0.05). Consequently, our findings strongly suggest that CNVs affecting SLCO3A1 may influence gene expression through a dosage effect. These results highlight the potential of SLCO3A1 as a candidate gene associated with aggressive traits in pig breeding programs.

Analysis of copy number variation in AZF region of Y chromosome in patients with spermatogenic failure

To explore the characteristics of copy number variation (CNV) within the Y chromosome azoospermia factor (AZF) region in patients with spermatogenesis disorders in the Shenzhen area. A total of 123 patients with spermatogenesis disorders who had visited Shenzhen People's Hospital from January 2016 to October 2022 (including 73 patients with azoospermia and 50 patients with oligozoospermia) and 100 normal semen males were selected as the study subjects. The AZF region was detected with multiplex ligation-dependent probe amplification (MLPA), and the correlation between the CNV in the AZF region and spermatogenesis disorders was analyzed using the chi-square test or Fisher's exact test. 19 CNV were detected among 53 patients from the 223 samples, including 20 cases (27.40%, 20/73) from the azoospermia group, 19 cases (38%, 19/50) from the oligozoospermia group, and 14 cases (14%, 14/100) from the normal control group. In the azoospermia, oligozoospermia, and normal control groups, the detection rates for CNV related to the AZFa region (including AZFab and AZFabc) were 5.48% (4/73), 2.00% (1/50), and 0 (0/100), respectively. The detection rates for the AZFb region (including the AZFbc region) were 6.85% (5/73), 0 (0/50), and 0 (0/100), respectively. The detection rates for gr/gr deletions in the AZFc region were 2.74% (2/73), 6.00% (3/50), and 9.00% (9/100), respectively, and those for b2/b4 deletions in the AZFc region were 2.74% (2/73), 10.00% (5/50), and 0 (0/100), respectively. The detection rates for complex rearrangements in the AZFc region were 6.85% (5/73), 18.00% (9/50), and 3.00% (3/100), respectively. Statistical analysis showed no significant difference in the detection rate of gr/gr deletions between the three groups (Fisher's Exact Test value = 2.712, P = 0.249); the differences in the detection rate of b2/b4 deletions between the three groups were statistically significant (Fisher's Exact Test value = 9.489, P = 0.002); the differences in the detection rate of complex rearrangements in the AZFc region between the three groups were statistically significant (Fisher's Exact Test value = 9.493, P = 0.006). In this study, a rare AZFa region ARSLP1 gene deletion (involving SY86 deletion) was detected in a patient with oligozoospermia. CNV in the AZFa and AZFb regions have a severe impact on spermatogenesis, but partial deletion in the AZFa region (ARSLP1 gene deletion) has a minor impact on spermatogenesis. The b2/b4 deletion and complex rearrangement in the AZFc region may be risk factors for male infertility. The gr/gr deletion may not serve as a risk factor for male infertility in the Shenzhen area.