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.
Stern
Rowena F
RF
Department of Medical and Molecular Genetics, Guy's, King's and St. Thomas' School of Medicine, London, UK. rowena.stern@genetics.kcl.ac.uk
Roberts
Roland G
RG
Mann
Kathy
K
Yau
Shu C
SC
Berg
Jonathan
J
Ogilvie
Caroline Mackie
CM
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