Detecting ligated fragments on oligonucleotide microarrays: optimizing chip design, array multiplex ligation-dependent probe amplification modification, and hybridization parameters.

Copy-number polymorphisms at specific genomic loci have been implicated in numerous human and animal disease phenotypes. Multiplex ligation-dependent probe amplification (MLPA) is a molecular genetic technique allowing targeted quantification of genomic copy-number changes (deletions and duplications), with potential for multiplexing up to 50 loci in one assay, and resolution down to the single nucleotide level. Modification of the MLPA technique to include Cy-labeled amplification primers permits parallel product detection by capillary electrophoresis and microarray hybridization. Detection and quantification of products by sequence-specific hybridization rather than size-specific capillary electrophoresis increases the potential for probe multiplexing possible in one assay and also allows for more flexible and efficient MLPA probe design. Protocols for the printing of synthetic oligonucleotide probe-sets for the detection of MLPA products, MLPA-probe amplification using array-compatible primers, and parallel product detection by quantitative capillary electrophoresis and microarray hybridization have been optimized.

The 12q14 microdeletion syndrome: six new cases confirming the role of HMGA2 in growth.

We report six patients with array deletions encompassing 12q14. Out of a total of 2538 array investigations carried out on children with developmental delay and dysmorphism in three diagnostic testing centres, six positive cases yielded a frequency of 1 in 423 for this deletion syndrome. The deleted region in each of the six cases overlaps significantly with previously reported cases with microdeletions of this region. The chromosomal range of the deletions extends from 12q13.3q15. In the current study, we report overlapping deletions of variable extent and size but primarily comprising chromosomal bands 12q13.3q14.1. Four of the six deletions were confirmed as de novo events. Two cases had deletions that included HMGA2, and both children had significant short stature. Neither case had osteopoikilosis despite both being deleted for LEMD3. Four cases had deletions that ended proximal to HMGA2 and all of these had much better growth. Five cases had congenital heart defects, including two with atrial septal defects, one each with pulmonary stenosis, sub-aortic stenosis and a patent ductus. Four cases had moderate delay, two had severe developmental delay and a further two had a diagnosis of autism. All six cases had significant speech delay with subtle facial dysmorphism.

Acute effects of C-peptide on gastric emptying in longstanding type 1 diabetes.

Gastric emptying (GE) of a solid (100 g beef) and liquid (150 ml 10 % dextrose) meal was measured in eight patients with type 1 diabetes during intravenous infusion of C-peptide (6 pmol/kg/ min) or isotonic saline. C-peptide had no effect on either solid or liquid GE.