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Use of two multiplex (PCR) reactions as an initial deletion screening method for DMD and BMD patients

Daisy Neves Falcão-Conceição^I; Márcia M. Gonçalves-Pimentel^I; Claudia P. Moreira^I; A.L.J. Kneppers^II; E. Bakker^II

^IDepartamento de Genética, Instituto de Biologia, Universidade Federal do Rio de Janeiro, Caixa Postal 68011, Ilha do Fundão, 21941 Rio de Janeiro, RI, Brasil. Send correspondence to D.N.F.-C.
^IIDepartment of Human Genetics, Leiden University, Leiden, The Netherlands

ABSTRACT

The dystrophin gene (located on Xp21) is an enormous gene-locus (more than 2.3 Mb). Mutations in this gene result in Duchene (DMD) or Becker (BMD) muscular dystrophy. About 60% of the mutations are deletions with an average size of 200 kb and are detectable by Southern blot analysis using cDNA probes. In 1988 Chamberlain et al. (Nucl. Acid. Res. 23: 11141-11156, 1988) described a multiplex reaction for a simultaneous amplification of nine exons from the dystrophin gene, to detect about 80% of all deletions. Recently Beggs et al. (Hum. Genet. 86:45-8, 1990) described an additional set of primers for a multiplex test of nine more exons.
We have used both multiplex reactions on 27 DMD and four BMD patients from Rio de Janeiro. Primers were synthesized and the kits were prepared in Leiden. With the Chamberlain multiplex reaction we detected 10 deletions. The second multiplex reaction (Beggs) confirmed seven of the deletions and detected two more deletions, one for exon 50 and another for exon 52. Southern blot and cDNA hybridization were used to confirm the deletions and to determine their extension. cDNA probes confirmed the 12 deletions detected using the two multiplex reactions. The multiplex approach as an initial screening for deletions is a good and reliable method.

Keywords: Multiplex PCR; DMD; BMD.

REFERENCES

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