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SALSA MLPA KIT P099B GCH1-TH
Autosomal dominant Dopa-responsive dystonia is characterized by postural and motor disturbances showing marked diurnal fluctuation (Segawa et al. (1976). The disorder is caused by a mutation in the gene encoding GTP cyclohydrolase I (GCH1). The GTP cyclohydrolase I enzyme is rate-limiting in the conversion of GTP to BH4, which is a cofactor for tyrosine hydroxylase. Tyrosine hydroxylase is the rate-limiting enzyme for dopamine synthesis. The GCH1 gene has 6 exons, spans 60 Kb of chromosomal sequence and is located on chromosome 14q22. The autosomal recessive form of dopa-responsive dystonia is also known as Segawa disease (DeLonlay P et al, 2000). Deficiency of Tyrosine Hydroxylase (TH) is generally considered as a cause of this autosomal recessive dystonia. TH is involved in the conversion of tyrosine to dopamine. As the rate-limiting enzyme in the synthesis of catecholamines, tyrosine hydroxylase has a key role in the physiology of adrenergic neurons. The TH gene is located in the Beckwith-Wiedemann syndrome (BWS) critical region (Gu et al, 2002) on chromosome 11p15.5. The TH gene has 14 exons and spans 8 Kb of chromosomal sequence. Mutations in the epsilon-sarcoglycan encoding SGCE gene cause myoclonus-dystonia syndrome. The SGCE gene has 11 exons and spans 72 Kb of chromosomal sequence on chromosome 7q21. Since the phenotypes of the dopa-responsive dystonia diseases are very similar, we combined probes for TH, GCH1 and SGCE exons in one MLPA probemix. The probemix contains probes for 5 of the 6 exons of GCH1 gene, for 5 of the 14 exons of TH gene and all 11 SGCE exons. As a control, 9 probes for other human genes located on different chromosomes are included. Unfortunately, it is not always possible to design a workable probe for each exon. Therefore, there is no probe present for exon 4 of GCH1 gene, nor are there probes included for exons 2, 5, 6, 7, 9, 10, 11 and 13 of the TH gene. However, all these exons are located in close proximity to neighboring exons. This MLPA kit is designed to detect deletions/duplications of one or more exons of the GCH1, TH, SGCE genes. Deletions of probe recognition sequences will be apparent by a 35-50% reduced relative peak area of the amplification product of that probe. However, mutations/polymorphisms very close to the probe ligation site may also result in a reduced relative peak area. Apparent deletions of a single exon therefore always require confirmation by other methods. We have no information on what percentage of defects in these genes is caused by deletions/duplications of complete exons. Please note that most defects in these genes are expected to be small (point) mutations, most of which will not be detected by this MLPA test.
Full mix description (pdf)

IMPORTANT NOTICE: MLPA kits are sold by MRC-Holland for research purposes and to demonstrate the possibilities of the MLPA technique. This kit is not CE/FDA certified for use in diagnostic procedures. Salsa MLPA kits are supplied with all necessary buffers and enzymes. Purchase of the Salsa MLPA test kits includes a limited license to use these products for research purposes. The use of this MLPA kit requires a thermocycler with heated lid and sequence type electrophoresis equipment. Different fluorescent PCR primers are available. The MLPA technique has been first described in Nucleic Acid Research 30, e57 (2002)
References 2007 -- Utility of MLPA in deletion analysis of GCH1 in dopa-responsive dystonia.
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