title: Multiplex PCR/liquid chromatography assay for screening of subtelomeric rearrangements.
abstract: In idiopathic or nonspecific mental retardation, the overall rate of cryptic subtelomeric rearrangements is estimated to be about 5%. Development of cost-effective screening for subtelomeric deletions would help clinical geneticists to make specific diagnoses in children with idiopathic mental retardation. Current screening modalities include fluorescence in situ hybridization (FISH) using subtelomeric probes and PCR-based quantitative analyses. Reductions in the cost and turnaround time will make the complete screening of subtelomeric rearrangements more widely used in clinical settings. Recently, a versatile method, called the multiplex PCR/liquid chromatography assay (MP/LC), was developed to assess copy numbers in this assay. Multiple genomic regions are amplified using unlabeled primers, then separated by ion-pair reversed-phase high-performance liquid chromatography. In the present study, we developed an MP/LC-based subtelomeric screening system that involves 21 multiple reactions and validated the protocol by analyzing 16 publicly available cell lines with known cytogenetic abnormalities involving at least one subtelomere per patient. To confirm the validity of the MP/LC method, we analyzed these cell lines concurrently with array-based comparative genomic hybridization (array-CGH), which gives higher resolution than the conventional G-banding technique. Among those 16 samples, the results from MP/LC and array-CGH agreed with each other perfectly. In 2 of the 16 samples, MP/LC correctly revealed subtelomeric duplications that were detected by array-CGH but were undetected by conventional cytogenetics, demonstrating the sensitivity of the MP/LC assay. This system is expected to be useful for making specific diagnoses and in genetic counseling for children with idiopathic mental retardation, a sizable fraction of whom have subtelomeric rearrangements.
authors.affiliation: Department of Pediatrics, Keio University School of Medicine, Tokyo, Japan.
authors.forename: Toru
authors.initials: T
authors.lastname: Udaka
authors.forename: Issei
authors.initials: I
authors.lastname: Imoto
authors.forename: Yoshinori
authors.initials: Y
authors.lastname: Aizu
authors.forename: Chiharu
authors.initials: C
authors.lastname: Torii
authors.forename: Kosuke
authors.initials: K
authors.lastname: Izumi
authors.forename: Rika
authors.initials: R
authors.lastname: Kosaki
authors.forename: Takao
authors.initials: T
authors.lastname: Takahashi
authors.forename: Shin
authors.initials: S
authors.lastname: Hayashi
authors.forename: Johji
authors.initials: J
authors.lastname: Inazawa
authors.forename: Kenjiro
authors.initials: K
authors.lastname: Kosaki
mesh_terms: D002460:Cell Line; D002853:Chromatography, Liquid / Q000379:methods*; D002869:Chromosome Aberrations; D002872:Chromosome Deletion; D017931:DNA Primers / Q000737:chemistry; D005820:Genetic Testing; D006801:Humans; D009693:Nucleic Acid Hybridization; D016133:Polymerase Chain Reaction / Q000379:methods*; D016615:Telomere / Q000235:genetics*
chemical_list: D017931:DNA Primers