Multiplex ligation-dependent probe amplification using a completely synthetic probe set

• Published in: BioTechniques Vol. 37; no. 3; pp. 399 - 405
• Main Authors: Stern, Rowena F, Roberts, Roland G, Mann, Kathy, Yau, Shu C, Berg, Jonathan, Ogilvie, Caroline Mackie
• Format: Journal Article
• Language: English
• Published: Natick, MA Future Science Ltd 01.09.2004; Eaton; Taylor & Francis Group
• Subjects: Biological and medical sciences; Chromosome Deletion; Diverse techniques; DNA Mutational Analysis - methods; DNA Probes - chemical synthesis; DNA Probes - genetics; Fundamental and applied biological sciences. Psychology; Gene Dosage; Molecular and cellular biology; Nucleic Acid Amplification Techniques - methods; Probe; Gene amplification; Ligation
• ISSN: 0736-6205; 1940-9818
• DOI: 10.2144/04373ST04

The recent development of multiplex ligation-dependent probe amplification (MLPA) has provided an efficient and reliable assay for dosage screening of multiple loci in a single reaction. However, a drawback to this method is the time-consuming process of generating a probe set by cloning in single-stranded bacteriophage vectors. We have developed a synthetic probe set to screen for deletions in a region spanning 18.5 Mb within chromosome 3q. In a pilot study, we tested 15 synthetic probes on 4 control samples and on 2 patients previously found to possess a heterozygous deletion in the region 3q26-q28. These synthetic probes detected deletions at all previously known deleted loci. Furthermore, using synthetic probes, the variability of results within samples was similar to that reported for commercially available M13-derived probes. Our results demonstrate that this novel approach to MLPA provides a generic solution to the difficulties of probe development by cloning; such synthetically generated probes may be used to screen a large number of loci in a single reaction. We conclude that the use of synthetic probes for MLPA is a rapid, robust, and efficient alternative for research (and potentially diagnostic) deletion and duplication screening of multiple genomic loci.