Multiplex ligation-dependent probe amplification using a completely synthetic probe set.
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.
Department of Medical and Molecular Genetics, Guy's, King's and St. Thomas' School of Medicine, London, UK. rowena.stern@genetics.kcl.ac.uk
Rowena F
RF
Stern
Roland G
RG
Roberts
Kathy
K
Mann
Shu C
SC
Yau
Jonathan
J
Berg
Caroline Mackie
CM
Ogilvie
D002872:Chromosome Deletion; D004252:DNA Mutational Analysis / Q000379:methods*; D015342:DNA Probes / Q000138:chemical synthesis* / Q000235:genetics; D018628:Gene Dosage; D021141:Nucleic Acid Amplification Techniques / Q000379:methods*
D015342:DNA Probes