title: Molecular characterization of ring chromosome 18 by low-coverage next generation sequencing.
abstract: BACKGROUND
  Ring chromosomes are one category of structurally abnormal chromosomes that can lead to severe growth retardation and other clinical defects. Traditionally, their diagnosis and characterization has largely relied on conventional cytogenetics and fluorescence in situ hybridization, array-based comparative genomic hybridization and single nucleotide polymorphism array-based comparative genomic hybridization. However, these methods are ineffectively at characterizing the ring chromosome structure and only offer a low resolution mapping of breakpoints. Here, we applied whole-genome low-coverage paired-end next generation sequencing (NGS) to two suspected cases of ring chromosome 18 (r(18)) and characterized the ring structure including the chromosome dosage changes and the breakpoint junction.
  
  
  METHODS
  The breakpoints and chromosome copy number variations (CNVs) of r(18) were characterized by whole-genome low-coverage paired-end NGS. We confirmed the dosage change by single nucleotide polymorphisms array, and validated the junction site regions using PCR followed by Sanger sequencing.
  
  
  RESULTS
  We successfully and fully characterized the r(18) in two cases by NGS. We mapped the breakpoints with a high resolution and identified all CNVs in both cases. We analyzed the breakpoint regions and discovered two breakpoints located within repetitive sequence regions, and two near the repetitive sequence regions. One of the breakpoints in case 2 was located within the gene METTL4, while the other breakpoints were intergenic.
  
  
  CONCLUSIONS
  We demonstrated that whole-genome low-coverage paired-end NGS can be used directly to map breakpoints with a high molecular resolution and detect all CNVs on r(18). This approach will provide new insights into the genotype-phenotype correlations on r(18) and the underlying mechanism of ring chromosomes formation. Our results also demonstrate that this can be a powerful approach for the diagnosis and characterization of ring chromosomes in the clinic.
authors.affiliation: State Key Laboratory of Reproductive Medicine, Department of Prenatal Diagnosis, Nanjing Maternity and Child Health Care Hospital Affiliated to Nanjing Medical University, Nanjing, China. xji0501@163.com.
authors.forename: Xiuqing
authors.initials: X
authors.lastname: Ji
authors.affiliation: State Key Laboratory of Reproductive Medicine, Department of Prenatal Diagnosis, Nanjing Maternity and Child Health Care Hospital Affiliated to Nanjing Medical University, Nanjing, China. liangdong2083@163.com.
authors.forename: Dong
authors.initials: D
authors.lastname: Liang
authors.affiliation: Department of Laboratory Medicine, The First Affiliated Hospital of Nanjing Medical University, Nanjing, China. sungirl515@163.com.|National Key Clinical Department of Laboratory Medicine, Nanjing, China. sungirl515@163.com.
authors.forename: Ruihong
authors.initials: R
authors.lastname: Sun
authors.affiliation: State Key Laboratory of Reproductive Medicine, Department of Prenatal Diagnosis, Nanjing Maternity and Child Health Care Hospital Affiliated to Nanjing Medical University, Nanjing, China. njfybjylcy@163.com.
authors.forename: Cuiyun
authors.initials: C
authors.lastname: Liu
authors.affiliation: State Key Laboratory of Reproductive Medicine, Department of Prenatal Diagnosis, Nanjing Maternity and Child Health Care Hospital Affiliated to Nanjing Medical University, Nanjing, China. dingyuanma2002@163.com.
authors.forename: Dingyuan
authors.initials: D
authors.lastname: Ma
authors.affiliation: State Key Laboratory of Reproductive Medicine, Department of Prenatal Diagnosis, Nanjing Maternity and Child Health Care Hospital Affiliated to Nanjing Medical University, Nanjing, China. njfybjywangyan@163.com.
authors.forename: Yan
authors.initials: Y
authors.lastname: Wang
authors.affiliation: State Key Laboratory of Reproductive Medicine, Department of Prenatal Diagnosis, Nanjing Maternity and Child Health Care Hospital Affiliated to Nanjing Medical University, Nanjing, China. njfybjyhuping@163.com.
authors.forename: Ping
authors.initials: P
authors.lastname: Hu
authors.affiliation: State Key Laboratory of Reproductive Medicine, Department of Prenatal Diagnosis, Nanjing Maternity and Child Health Care Hospital Affiliated to Nanjing Medical University, Nanjing, China. zhengfeng_xu_nj@163.com.
authors.forename: Zhengfeng
authors.initials: Z
authors.lastname: Xu
mesh_terms: D001483:Base Sequence; D002874:Chromosome Mapping; D002887:Chromosomes, Human, Pair 18 / Q000235:genetics; D019295:Computational Biology; D020732:Cytogenetic Analysis; D005260:Female; D005333:Fetus; D018628:Gene Dosage / Q000235:genetics*; D059014:High-Throughput Nucleotide Sequencing / Q000379:methods; D006801:Humans; D008780:Methyltransferases / Q000235:genetics; D008969:Molecular Sequence Data; D010641:Phenotype; D016133:Polymerase Chain Reaction; D020641:Polymorphism, Single Nucleotide / Q000235:genetics; D011296:Prenatal Diagnosis; D012303:Ring Chromosomes; D017422:Sequence Analysis, DNA
chemical_list: C586526:METTL4 protein, human; D008780:Methyltransferases