2008 Barcelona - European Society of Human Genetics

Normal variation, population genetics, genetic epidemiology
deletions (Indels) . Nucleotide substitutions were found to make up the
majority of the mutations, rather than indels. We drew significantly
high transition rate (81 .8%) versus lower frequency of transversions
(18.2%). 12 polymorphisms were identified in this study which had not
been published in the MitoMap database . PCT changes at position
303-309 were detected in 83% of our samples . Our results suggest
that an increased level of HVS-I and HVS-II substitutions may be an
indicator of mitochondrial DNA instability . Furthermore, mtDNA mutations
may play an important role in pathogenesis of cardiac arrest
which has remained unexplained for long .
P07.066
Frequency of the Hemochromatosis gene mutations in patients
with Hereditary Hemochromatosis and in control subjects from
serbia
M. Saric1 , L. Zamurovic1 , M. Keckarevic-Markovic1 , D. Keckarevic1 , M. Kecmanovic1
, D. Savic-Pavicevic1 , J. Jovic2 , S. Romac1 ;
1 2 Faculty of Biology, Belgrade, Serbia, Military Medical Academy, Belgrade,
Serbia.
The most common form of hereditary hemochromatosis (HH) is HH
type 1 . It is an autosomal recessive iron-overload disorder associated
with mutations in the hemochromatosis gene (HFE) . The aim of this
study was to determine the frequency of the C282Y, H63D and S65C
mutations of HFE gene in the group of unrelated HH patients (n=28)
and in the healthy population of Serbia (n=318) .
The C282Y, H63D and S65C mutation frequencies in the healthy population
were 1 .6%, 15 .7% and 1 .6%, respectively and in the population
of HH patients 25%, 19 .6% and 1 .8%, respectively . The frequency
of C282Y homozygotes was estimated on approximately 1:3900 indicating
the prevalence of HH type 1 in Serbia . Our results support
previously documented gradient distribution of the C282Y mutation, in
northwest to southeast Europe. They also confirm that the frequency
of C282Y mutation is not related to the origin of particular population
but it is more related to the geographical position of the population .
This finding could be considered as yet further evidence in favor of the
Viking or Celtic origin of C282Y mutation .
The frequency of the C282Y homozygotes (10 .7%) is considerably
lower than the frequency reported for HH patients in other European
populations . It is possible that some of the analyzed HH patients carry
other mutations in HFE gene, mutation in some other genes already
related with HH or in some other yet not identified genes.
These results have important clinical implications for the detection and
management of HH type 1 in Serbia .
P07.067
Hereditary hemochromatosis and multiple sclerosis. Frequency
of common HFE mutations in czech patients with multiple
sclerosis
M. Tomková1 , P. Strachotová1 , L. Barnincová1 , E. Krasulová1 , L. Pospíšilová1 ,
D. Horáková1 , E. Havrdová1 , J. Horák2 , P. Martásek1 ;
1 2 1st School of Medicine, Prague, Czech Republic, 3rd School of Medicine,
Prague, Czech Republic.
Hereditary hemochromatosis is an autosomal recessive disorder frequently
found in Caucasian population and caused by iron overload
(mutations in HFE gene) . There have been several reports of a relationship
between HFE gene polymorphisms and multiple sclerosis (MS) .
MS is an inflammatory demyelinating disease of the central nervous
system . In the etiology of MS, both environmental and genetic factors
play a role . In this study we investigated whether HFE polymorphisms
is associated with Czech MS patients . We therefore genotyped 410 MS
patients and 481 healthy controls for the C282Y, H63D, and S65C mutations
by PCR-RFLP analysis. None of the MS patients were identified
as homozygous for S65C or C282Y mutation, and seven (1 .70%) for
H63D mutation . Twenty eight (6 .83%) MS patients were C282Y heterozygous,
seventy seven (18 .78%) were H63D heterozygous, and eight
(1 .95%) were S65C heterozygous . Among them, we have found four
(0 .97%) compound heterozygotes; three patients were C282Y/H63D
compound heterozygous and one was found H63D/S65C compound
heterozygous. Our results showed no significant differences in the distribution
of C282Y and S65C HFE mutations between MS patients and
controls . However, the allele frequency of the H63D mutation was significantly
lower (p < 0,05) in the MS patients than in the control group.
(Supported by grant IGA MZ 1A8713 and MSM 0021620806) .
P07.068
comparison of genome-wide homozygosity-by-descent
estimates in human isolated population
O. Polasek 1,2 , I. Kolčić 2 , L. Zgaga 2 , Z. Biloglav 2 , A. Vorko Jović 2 , D. Rudan 3 , I.
Rudan 4 , A. Wright 5 , H. Campbell 1 , A. Leutenegger 6,7 ;
1 Public Health Sciences, University of Edinburgh, Edinburgh, United Kingdom,
2 Andrija Stampar School of Public Health, Medical School, University of Zagreb,
Zagreb, Croatia, 3 Clinical Hospital Dubrava, Zagreb, Croatia, 4 Centre for
Global Health, University of Split, Split, Croatia, 5 Human Genetics Unit, Medical
Research Council, Edinburgh, United Kingdom, 6 Inserm U535, Villejuif, France,
7 Univ. Paris-Sud, Villejuif, France.
The aim of this study was to investigate several methods of marker
based genome-wide homozygosity estimation . The study sample
consisted of 118 members of a single extended family living on the
Adriatic Croatian island of Vis, where no pedigree based inbreeding
was observed, and therefore no homozygosity by descent, HBD, was
expected . Individuals were genotyped for 810 microsatellite markers
and a 317k Illumina chip . Five approaches were compared: the proportion
of heterozygous loci (multilocus heterozygosity, MLH), two methods
of moments approaches that use different weighting approaches
(ADC and PLINK), and finally 2 maximum likelihood approaches, one
singlepoint and the other multipoint, using a hidden Markov model for
marker dependencies (FEstim) . The latter is the only existing multipoint
method for HBD estimation and gives estimates the closest to
the true HBD values, as shown previously in a simulation study . The
results indicated that the two methods of moments approaches correlated
highly with MLH and hence did not bring much information about
HBD . On the other hand, the two maximum likelihood approaches
exhibited zero value estimates when homozygosity was likely due to
chance. A total of 68 individuals (57.6%) had no HBD as defined by the
FEstim, while a total of 4 individuals (3 .9%) had FEstim values over
0.0625 indicating inbreeding closer than first cousins. These methods
can be used to differentiate homozygosity by chance vs . HBD, which
may be important in investigation of the genome-wide heterozygosity
effects on quantitative and health related traits .
P07.069
A medium-throughput assay for the genotyping and primary
screening of useful polymorphisms using High Resolution
melting analysis
X. Muñoz, F. Marín, N. García, N. Sala;
Catalan Institute of Oncology - Translational Research Laboratory and Unit of
Nutrition, Environment and Cancer, L’Hospitalet de Llobregat, Spain.
The High Resolution Melting (HRM) analysis uses highly sensitive fluorescent
dyes specific for double stranded DNA and the Post-PCR melting
curve, to characterize gene fragments and identify small sequence
variants such as SNPs . The purpose of this study was to determine the
performance of LightCycler® 480 High Resolution Melting master and
the LightCycler® 480 Gene Scanning Software, for genotyping and
gene scanning purposes in the LightCycler® 480 (LC480) System . For
this we setup protocols for the analysis of the MTHFR c .677C>T, FII
c .20210G>A, and F5 Leiden variants in 60 samples previously genotyped
using hybridization probes . Among these samples 20 there were
heterozygotes for MTHFR c .677C>T, 10 for FII c .20210G>A and 4 for
the F5 Leiden variant . The remaining samples were wild-type homozygotes
We have also developed a protocol for the mutational scanning
of Trefoil factor 2 gene (TFF2) . Post PCR HRM resulted in the expected
different melting profiles that efficiently discriminated between homozygotes
and heterozygotes for the different polymorphisms tested in
all samples . HRM scanning of TFF2 exon 4 in 50 control samples from
our population, allowed for the identification of a previously described
SNP (rs225334; MAF: 0 .39) and a novel insertion, c .68_69insCTT
(MAF: 0 .05) in the 3’-UTR region . Both variants could be genotyped
simultaneously in the same amplicon melting analysis .
We conclude that HRM may be an efficient and cost-effective method
for the simultaneous amplification, mutational scanning and genotyping
of amplicons up to 200bp, in a single step and in sets of 96 or 384
samples .