Simultaneous sequencing of 37 genes identified causative mutations in the majority of children with renal tubulopathies.

The clinical diagnosis of inherited renal tubulopathies can be challenging as they are rare and characterized by significant phenotypic variability. Advances in sequencing technologies facilitate the establishment of a molecular diagnosis. Therefore, we determined the diagnostic yield of a next generation sequencing panel assessing relevant disease genes in children followed through three national networks with a clinical diagnosis of a renal tubulopathy.

Conformation-sensitive capillary electrophoresis.

Conformation-sensitive capillary electrophoresis (CSCE) is a rapid, high-throughput screening method that can be applied to any region of a genome for detection of sequence variants. Slab gel-based conformation-sensitive gel electrophoresis was first described by Ganguly et al., and the transfer from slab gels to capillaries for higher throughput was reported by Rozycka et al.

Marked hemiatrophy in carriers of Duchenne muscular dystrophy.

Objective: To describe the clinical and molecular genetic findings in 2 carriers of Duchenne muscular dystrophy (DMD) who exhibited marked hemiatrophy. Duchenne muscular dystrophy is an X-linked disorder in which affected male patients harbor mutations in the dystrophin gene. Female patients with heterozygous mutations may be manifesting carriers.

Simultaneous mutation scanning for gross deletions, duplications and point mutations in the DMD gene.

We have developed a technique to screen for gross deletions/duplications and point mutations using one streamlined approach. Fluorescent multiplex quantitative PCR is used to determine the copy number of each exon, followed by conformation sensitive capillary electrophoresis (CSCE) of the same PCR products on a multi-capillary genetic analyser. We have developed this technique to screen all 79 exons of one of the largest human genes currently known (dystrophin) using 12 multiplex PCR assays.

Simultaneous MLPA-based multiplex point mutation and deletion analysis of the dystrophin gene.

The Multiplex Ligation-dependent Probe Amplification assay (MLPA) is the method of choice for the initial mutation screen in the analysis of a large number of genes where partial or total gene deletion is part of the mutation spectrum. Although MLPA dosage probes are usually designed to bind to normal DNA sequence to identify dosage imbalance, point mutation-specific MLPA probes can also be made. Using the dystrophin gene as a model, we have designed two MLPA probe multiplexes that are specific to a number of commonly listed point mutations in the Leiden dystrophin point mutation database (http://www.