title: Array-MLPA: comprehensive detection of deletions and duplications and its application to DMD patients.
abstract: Multiplex ligation-dependent probe amplification (MLPA) is widely used to screen genes of interest for deletions and duplications. Since MLPA is usually based on size-separation of the amplification products, the maximum number of target sequences that can be screened in parallel is usually limited to approximately 40. We report the design of a robust array-based MLPA format that uses amplification products of essentially uniform size (100-120 bp) and distinguishes between them by virtue of incorporated tag sequences. We were thus able to increase probe complexity to 124, with very uniform product yields and signals that have a low coefficient of variance. The assay designed was used to screen the largest set studied so far (249 patients) of unrelated Duchenne muscular dystrophy (DMD) cases from the Chinese population. In a blind study we correctly assigned 98% of the genotypes and detected rearrangements in 181 cases (73%); i.e., 163 deletions (65%), 13 duplications (5%), and five complex rearrangements (2%). Although this value is significantly higher for Chinese patients than previously reported, it is similar to that found for other populations. The location of the rearrangements (76% in the major deletion hotspot) is also in agreement with other findings. The 96-well flow-through microarray system used in this research provides high-throughput and speed; hybridization can be completed in 5 to 30 minutes. Since array processing and data analysis are fully automated, array-MLPA should be easy to implement in a standard diagnostic laboratory. The universal array can be used to analyze any tag-modified MLPA probe set.
authors.affiliation: Shanghai Institute of Medical Genetics, Shanghai Children's Hospital, Shanghai Jiao Tong University School of Medicine, PR China.
authors.forename: Fanyi
authors.initials: F
authors.lastname: Zeng
authors.forename: Zhao-Rui
authors.initials: ZR
authors.lastname: Ren
authors.forename: Shang-Zhi
authors.initials: SZ
authors.lastname: Huang
authors.forename: Margot
authors.initials: M
authors.lastname: Kalf
authors.forename: Monique
authors.initials: M
authors.lastname: Mommersteeg
authors.forename: Maarten
authors.initials: M
authors.lastname: Smit
authors.forename: Stefan
authors.initials: S
authors.lastname: White
authors.forename: Chun-Lian
authors.initials: CL
authors.lastname: Jin
authors.forename: Miao
authors.initials: M
authors.lastname: Xu
authors.forename: Da-Wen
authors.initials: DW
authors.lastname: Zhou
authors.forename: Jing-Bin
authors.initials: JB
authors.lastname: Yan
authors.forename: Mei-Jue
authors.initials: MJ
authors.lastname: Chen
authors.forename: Rinie
authors.initials: R
authors.lastname: van Beuningen
authors.forename: Shu-Zhen
authors.initials: SZ
authors.lastname: Huang
authors.forename: Johan
authors.initials: J
authors.lastname: den Dunnen
authors.forename: Yi-Tao
authors.initials: YT
authors.lastname: Zeng
authors.forename: Ying
authors.initials: Y
authors.lastname: Wu
mesh_terms: D005260:Female; D017353:Gene Deletion; D020440:Gene Duplication; D005820:Genetic Testing / Q000379:methods*; D006801:Humans; D008297:Male; D020388:Muscular Dystrophy, Duchenne / Q000175:diagnosis* / Q000235:genetics*; D021141:Nucleic Acid Amplification Techniques / Q000379:methods*