Rapid detection of subtelomeric deletion/duplication by novel real-time quantitative PCR using SYBR-green dye

Hum Mutat. 2004 Apr;23(4):368-78. doi: 10.1002/humu.20011.


Telomeric chromosome rearrangements may cause mental retardation, congenital anomalies, miscarriages, and hematological malignancies. Automated detection of subtle deletions and duplications involving telomeres is essential for high-throughput screening procedures, but impractical when conventional cytogenetic methods are used. Novel real-time PCR quantitative genotyping of subtelomeric amplicons using SYBR-green dye allows high-resolution screening of single copy number gains and losses by their relative quantification against a diploid genome. To assess the applicability of the technique in the screening and diagnosis of subtelomeric imbalances, we describe here a blinded study in which DNA from 20 negative controls and 20 patients with known unbalanced cytogenetic abnormalities involving at least one or more telomeres were analyzed using a novel human subtelomere-specific primer set, producing altogether 86 amplicons, in the SYBR-green I-based real-time quantitative PCR screening approach. Screening of the DNA samples from 20 unrelated controls for copy number polymorphism do not detect any polymorphism in the set of amplicons, but single-copy-number gains and losses were accurately detected by quantitative PCR in all patients, except the copy number alterations of the subtelomeric p-arms of the acrocentric chromosomes in two cases. Furthermore, a detailed mapping of the deletion/translocation breakpoint was demonstrated in two cases by novel real-time PCR "primer-jumping." Because of the simplicity and flexibility of the SYBR-green I-based real-time detection, the primer-set can easily be extended, either to perform further detailed molecular characterization of breakpoints or to include amplicons for the detection and/or analysis of syndromes that are associated with genomic copy number alterations, e.g., deletion/duplication-syndromes and malignant cancers.

Publication types

  • Evaluation Study

MeSH terms

  • Benzothiazoles
  • Chromosome Aberrations*
  • Chromosome Breakage
  • Chromosome Disorders / diagnosis*
  • Chromosome Disorders / genetics
  • Chromosomes, Human
  • Diamines
  • Female
  • Fluorescent Dyes*
  • Humans
  • Male
  • Organic Chemicals*
  • Polymerase Chain Reaction / methods*
  • Quinolines
  • Sequence Deletion
  • Telomere / genetics*
  • Time Factors
  • Translocation, Genetic


  • Benzothiazoles
  • Diamines
  • Fluorescent Dyes
  • Organic Chemicals
  • Quinolines
  • SYBR Green I