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PGD for Primary dystonia
A multiplex fluorescent PCR protocol, including simultaneous
testing of the DYT1 gene mutation and a set of linked
polymorphic markers, is used for early-onset primary
dystonia (PD), an autosomal dominant disorder caused, in
most of the cases, by a 3bp deletion (codon 302) in the DYT1
gene. Mutation analysis was performed simply by sizing PCR
fragments, detecting the affected alleles because of the 3bp
difference from the normal allele (Figure 2).
Because this
mutation is responsible for more than 70% of PD cases, the PGD strategy used can be applied for most of patients
without the need for extensive preclinical preparatory work.
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Figure 2.
Preimplantation genetic diagnosis (PGD) for Primary
dystonia (PD).
A) Haplotyping results of STR markers linked to DYT1
gene are numbered according to the size in base pair
(bp). Numbers in bold represent alleles linked to
the mutation. Examples of different results of DYT1
mutation analysis from biopsied blastomeres are
shown in the lower panel.
B) Capillary electrophoresis of fluorescent
polymerase chain reaction (PCR) products obtained
after multiplex amplification of the DYT1 region
involved by mutation and a set of linked polymorphic
markers.
The x-axis shows the length of PCR products
in bp and the y-axis shows the fluorescence
intensity in Relative Fluorescence Units (RFU).
Mutation analysis was performed by determining the
size of the PCR fragments, detecting the affected
alleles (113 bp peak) because of the 3bp difference
from the normal allele (116 bp peak).
On top of the electropherogram the marker name is located above
the corresponding alleles (peaks). Numbers next to
each peak represent the size of the allele (in bp).
Alleles linked to the mutation are highlighted in
green.
The upper lane shows the results of an
affected embryo (embryo 1). An affected embryo
(embryo 2) in which the mutated allele (113 bp peak)
of DYT1 gene has dropped out is shown in the middle
lane. Allele drop-out occurrence has been evidenced
from the presence of the affected haplotype
(highlighted peaks).
The lower lane shows the
profile of a healthy embryo (embryo 3). |
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PGD for Duchenne muscular dystrophy (DMD)
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