Analysis of 17p11.2 chromosome region rearrangements in CMT1 patients from Ukraine
Two intercomplementary methods of 17p11.2 duplication/deletion identification have been elaborated: STR allelic variants analysis and direct PMP22 gene dosage measuring by means of quantitative Real Time PCR. It has been carried out detection and analysis of 17p11.2 chromosome region rearrangements...
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Hryshchenko, N.V. Livshits, L.A. 2014-07-19T12:11:53Z 2014-07-19T12:11:53Z 2009 Analysis of 17p11.2 chromosome region rearrangements in CMT1 patients from Ukraine / N.V. Hryshchenko, L.A. Livshits // Цитология и генетика. — 2009. — Т. 43, № 1. — С. 36-41. — Бібліогр.: 14 назв. — англ. 0564-3783 https://nasplib.isofts.kiev.ua/handle/123456789/66620 575.11+577.21+577.213.3+616.832–009.55 Two intercomplementary methods of 17p11.2 duplication/deletion identification have been elaborated: STR allelic variants analysis and direct PMP22 gene dosage measuring by means of quantitative Real Time PCR. It has been carried out detection and analysis of 17p11.2 chromosome region rearrangements in CMT1 patients from Ukraine. It has been registered the high level of de novo cases with 17p11.2-duplication. It has been shown the 17p11.2 chromosome region duplication/deletion association with CMT1A and HNPP clinical phenotypes which may be used in differential diagnosis of this type of CMT polyneuropathy. Проведена детекция и анализ хромосомных перестроек области 17p11.2 у пациентов с болезнью ШаркоМари-Тус (ШМТ) из Украины. Разработаны два взаимодополняющих метода идентификации дупликаций/делеций в области 17p11.2: анализ аллельных вариантов STR локусов и прямой анализ дозы гена РМР22 с использованием ПЦР в реальном времени. Выявлен высокий уровень de novo дупликаций в указанном хромосомном регионе. Показана ассоциация дупликаций/делеций в области 17p11.2 с клиническими фенотипами ШМТ1А и HNPP, что может быть использовано для дифференциальной диагностики упомянутых типов полиневропатии ШМТ. Проведено детекцію та аналіз хромосомних перебудов ділянки 17p11.2 у пацієнтів з хворобою Шарко-Марі-Тус (ШМТ) з України. Розроблено два взаємодоповнюючі методи ідентифікації дуплікацій/делецій у ділянці 17p11.2: аналіз алельних варіантів STR-локусів та прямий аналіз дози гена РМР22 із використанням ПЛР у реальному часі. Виявлено високий рівень de novo дуплікацій цієї хромосомної ділянки. Показано ассоціацію дуплікацій/делецій у ділянці 17p11.2 із клінічними фенотипами ШМТ1А та HNPP, що може бути використано для диференційної діагностики даних типів поліневропатії ШМТ. en Інститут клітинної біології та генетичної інженерії НАН України Цитология и генетика Оригинальные работы Analysis of 17p11.2 chromosome region rearrangements in CMT1 patients from Ukraine Анализ хромосомных перестроек в области 17р11.2 у больных ШМТ1 из Украины Аналіз хромосомних перебудов у ділянці 17р11.2 у хворих ШМТ1 з України Article published earlier |
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| title |
Analysis of 17p11.2 chromosome region rearrangements in CMT1 patients from Ukraine |
| spellingShingle |
Analysis of 17p11.2 chromosome region rearrangements in CMT1 patients from Ukraine Hryshchenko, N.V. Livshits, L.A. Оригинальные работы |
| title_short |
Analysis of 17p11.2 chromosome region rearrangements in CMT1 patients from Ukraine |
| title_full |
Analysis of 17p11.2 chromosome region rearrangements in CMT1 patients from Ukraine |
| title_fullStr |
Analysis of 17p11.2 chromosome region rearrangements in CMT1 patients from Ukraine |
| title_full_unstemmed |
Analysis of 17p11.2 chromosome region rearrangements in CMT1 patients from Ukraine |
| title_sort |
analysis of 17p11.2 chromosome region rearrangements in cmt1 patients from ukraine |
| author |
Hryshchenko, N.V. Livshits, L.A. |
| author_facet |
Hryshchenko, N.V. Livshits, L.A. |
| topic |
Оригинальные работы |
| topic_facet |
Оригинальные работы |
| publishDate |
2009 |
| language |
English |
| container_title |
Цитология и генетика |
| publisher |
Інститут клітинної біології та генетичної інженерії НАН України |
| format |
Article |
| title_alt |
Анализ хромосомных перестроек в области 17р11.2 у больных ШМТ1 из Украины Аналіз хромосомних перебудов у ділянці 17р11.2 у хворих ШМТ1 з України |
| description |
Two intercomplementary methods of 17p11.2 duplication/deletion identification have been elaborated: STR allelic variants analysis and direct PMP22 gene dosage measuring by means of quantitative Real Time PCR. It has been carried out detection and analysis of 17p11.2 chromosome region rearrangements in CMT1 patients from Ukraine. It has been registered the high level of de novo cases with 17p11.2-duplication. It has been shown the 17p11.2 chromosome region duplication/deletion association with CMT1A and HNPP clinical phenotypes which may be used in differential diagnosis of this type of CMT polyneuropathy.
Проведена детекция и анализ хромосомных перестроек области 17p11.2 у пациентов с болезнью ШаркоМари-Тус (ШМТ) из Украины. Разработаны два взаимодополняющих метода идентификации дупликаций/делеций в области 17p11.2: анализ аллельных вариантов STR локусов и прямой анализ дозы гена РМР22 с использованием ПЦР в реальном времени. Выявлен высокий уровень de novo дупликаций в указанном хромосомном регионе. Показана ассоциация дупликаций/делеций в области 17p11.2 с клиническими фенотипами ШМТ1А и HNPP, что может быть использовано для дифференциальной диагностики упомянутых типов полиневропатии ШМТ.
Проведено детекцію та аналіз хромосомних перебудов ділянки 17p11.2 у пацієнтів з хворобою Шарко-Марі-Тус (ШМТ) з України. Розроблено два взаємодоповнюючі методи ідентифікації дуплікацій/делецій у ділянці 17p11.2: аналіз алельних варіантів STR-локусів та прямий аналіз дози гена РМР22 із використанням ПЛР у реальному часі. Виявлено високий рівень de novo дуплікацій цієї хромосомної ділянки. Показано ассоціацію дуплікацій/делецій у ділянці 17p11.2 із клінічними фенотипами ШМТ1А та HNPP, що може бути використано для диференційної діагностики даних типів поліневропатії ШМТ.
|
| issn |
0564-3783 |
| url |
https://nasplib.isofts.kiev.ua/handle/123456789/66620 |
| citation_txt |
Analysis of 17p11.2 chromosome region rearrangements in CMT1 patients from Ukraine / N.V. Hryshchenko, L.A. Livshits // Цитология и генетика. — 2009. — Т. 43, № 1. — С. 36-41. — Бібліогр.: 14 назв. — англ. |
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2025-11-26T11:31:02Z |
| last_indexed |
2025-11-26T11:31:02Z |
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| fulltext |
УДК 575.11+577.21+577.213.3+616.832–009.55
N.V. HRYSHCHENKO, L.A. LIVSHITS
Institute of Molecular Biology and Genetics, NASU, 150 Zabolotnogo str.,
Kyiv, Ukraine, 03680, livshits@imbmg.org.ua
ANALYSIS OF 17p11.2
CHROMOSOME REGION
REARRANGEMENTS IN CMT1
PATIENTS FROM UKRAINE
Two intercomplementary methods of 17p11.2 duplica�
tion/deletion identification have been elaborated: STR allelic
variants analysis and direct PMP22 gene dosage measuring
by means of quantitative Real�Time PCR. It has been carried
out detection and analysis of 17p11.2 chromosome region
rearrangements in CMT1 patients from Ukraine. It has been
registered the high level of de novo cases with 17p11.2�duplica�
tion. It has been shown the 17p11.2 chromosome region dupli�
cation/deletion association with CMT1A and HNPP clinical
phenotypes which may be used in differential diagnosis of this
type of CMT polyneuropathy.
Introduction. Charcot–Marie–Tooth disease
(CMT) and related neuropathies (hereditary neu�
ropathy with liability to pressure palsies [HNPP]
and Dejerine�Sottas disease [DSS]) are clinically
and genetically heterogeneous group of sensorineur�
al polyneuropathies. CMT hereditary neuropathy
syndrome is the most common genetic cause of
neuropathy with a frequency of 1 in 2500 [1].
The first level of classification concerns the
mode of inheritance: autosomal dominant, X�linked
and autosomal recessive. The second level of clas�
sification in each subgroup depends upon the neu�
ropathy disorder process. CMT is traditionally
classified into two types, demyelinating (CMT1)
and axonal (CMT2) forms according to electro�
physiological criteria The diagnostic criteria for
Charcot�Marie�Tooth disease type 1 are motor
nerve conduction velocities under 38 m/s, on ion
bulbs and signs of demyelination and remyelina�
tion in sural nerve biopsies, and no clinical or
histopathological evidence of other diseases asso�
ciated with demyelinating polyneuropathies.
CMT2 is characterized by normal or slightly
reduced motor nerve conduction velocities [2].
Many loci and genes have been identified in
each CMT subtype. Autosomal dominant demyeli�
nating CMT can be due mostly to PMP22 gene
17p11.2�duplication (CMT1A) but also to other
genes such as P0 (1q22), LITAF (16p13) and
EGR2 (10q21) mutations. Approximately 20 % of
all individuals presenting neuromuscular clinics
with a chronic peripheral neuropathy have CMT
type 1A (PMP22 duplication) as the cause of the
neuropathy [3]. Prevalence is about 1 in 3300 peo�
ple. The uncommon allelic variant of CMT1A is
Herditary Neuropathy with Liability to Pressure
Palsies (HNPP).
First described by De Jong in 1947 HNNP is
an autosomal dominant inherited disease charac�
terized by recurrent painless focal neuropathies.
Chance et al identified a 1.5Mb reciprocal deletion
encompassing the PMP22 gene which is responsi�
ble for the disease [4]. The same region (17p11.2)
duplication leads to distinct clinical type –
CMT1A. The CMT1A duplication and HNPP
deletion represent products of unequal crossing
over and a reciprocal recombination between
flanking 24�kb homologous sequences termed
CMT1A–REPs [5, 6] (Fig. 1).
The differential DNA�diagnostic of CMT1A/
HNPP is based on the 17p11.2 duplication/dele�
tion or PMP22 gene point mutation detection.
ISSN 0564–3783. Цитология и генетика. 2009. № 136
© N.V. HRYSHCHENKO, L.A. LIVSHITS, 2009
The methods used in diagnostic laboratories to
detect the 17p11.2 duplication/deletion include
quantitative methods and indirect methods such as
pulsed field gel electrophoresis, microsatellite
analysis, detection of the common junction frag�
ment by Southern blotting, detection of the com�
mon junction fragment by polymerase chain reac�
tion (PCR), and dosage analysis of PMP�22 using
semi�quantitative fluorescent PCR (STS dosage)
with products analysed using either capillary elec�
trophoresis or polyacrylamide gel electrophoresis
(PAGE) and finally quantitative Real�Time dosage
analysis of PMP�22 gene [7–11].
In the present study we have provide detection
and analysis of 17p11.2 chromosome region
rearrangements in CMT1 patients from Ukraine
using SRT�PCR method combined with real�time
quantitative PCR.
Materials and Methods. The subjects were DNA�
samples extracted from peripheral blood leuco�
cytes from Ukrainian CMT�patients and their rel�
atives (total 57 CMT families with dominant and
unclear inheritance). For the current purpose we
exclude those CMT�families who revealed recessive
type of inheritance. In our investigation DNA
samples from unrelated volunteer non�CMT donors
from different regions of Ukraine were used as
normal controls. Informed consents were obtained
from all individuals participating in our study.
DNA isolation from blood samples containing
glugicir anticoagulant were done by standard phe�
nol/chloroform procedures [12]. Quantity and
quality of DNA probes were analyzed on ND�
1000 spectrophotometer (NanoDrop, USA).
STR�PCR method of CMT1A/HNPP detection.
The STR loci D17S122, D17S921, D17S1358 and
D17S2226 from 17р11.2 chromosomal region were
used for the current study. Primer design, PCR
conditions and fragment analysis using automated
DNA «ALF express II» sequencer (Amersham Bio�
science) of the three (CA) n STR loci allelic vari�
ants were optimized and provided as were pub�
lished previously [13]. D17S2226 tetranucleotide
polymorphism alleles were amplified using pub�
lished primer sequences [7] and analyzed the same
way as (CA) n STR loci.
Real�Time quantitative PCR method of CMT1A/
HNPP detection. For PMP22 exon 3 (target
sequence) and ALB exon 12 (reference sequence)
amplification we have used previously published
ІSSN 0564–3783. Цитология и генетика. 2009. № 1 37
Analysis of 17p11.2 chromosome region rearrangements in CMT1 patients from Ukraine
Fig. 1. 17p11.2 chromosome region rearrangements
Fig. 2. Duplication and deletion detection in 17p11.2 region:
a – case with CMT1A�duplication ALF�express fluoro�
gramm. D17S1358 locus: 1 – father, 2 – son (inherited
maternal chromosome without duplication), 3 – mother
(has CMT1A duplication); b – case with HNPP�deletion
ALF�express fluorogramm; 1, 2 – D17S1358 locus, 3, 4 –
D17S921 locus, 5, 6 – D17S2226 locus, 1, 3, 5 – son (has
paternal chromosome with deletion), 2, 4, 6 – father with
deletion of 17p11.2 region
primers [11]. A 4�by�4�primer matrix (combina�
tions of 5 μM, 10 μM, 20 μM and 40 μM of each
forward and reverse primers) was analyzed to
determine the optimal concentrations of both for�
ward and reverse primers. The minimum primer
concentrations that resulted in the lowest Ct�value
(threshold cycle) and highest fluorescent signal
(�Rn), while minimizing nonspecific amplifica�
tion, were chosen as an optimal pair. The amplifi�
cations were carried out in a 50 μl reaction volume
containing 25 μl iQTM SYBR Green Supermix
Sample (BIO�RAD, USA), 10 μM forward primer
and reverse primer, and 20 ng (no less then 4 μl) of
genomic DNA. In each PCR, normal controls
(non�CMT) and no�template control were includ�
ed. Each sample was run in duplicate for both
PMP22 and ALB sequences amplification. PCR
reactions were run in the iQ5TM Multicolor Real�
Time PCR Detection System (BIO�RAD, USA).
Preincubation was performed for 5 min at 95 °С to
denature the target DNA and activate hot�start
iTaqTM DNA polymerase. DNA samples were
amplified for 40 cycles of 15 sec at 95 °С and
1 min at 65 °С. The fluorescence signals were
measured in Real�Time mode at the end of the
elongation phase. Amplicons were run as dupli�
cates in separate tubes to permit quantification of
the PMP22 gene normalized to ALB gene (albu�
min), an endogenous gene, as a control. By using a
calibrator sample of normal control DNA with
2 copies of PMP22 (verified by STR�PCR
method), the relative gene copy number (��Ct
ratio) of unknown sample was estimated using a
2–��Ct method (Livak), because of both target and
reference genes appear to be amplified with effi�
ciencies near 100 % and within 5 % of each other
(data not shown). The Real�Time data were pro�
cessing by use the BIO�RAD iQ5 Optical System
Software V 2.0 (2006).
Results and Discussion. As was mentioned
above several methods have been developed in
clinical laboratories for the molecular diagnosis of
CMT1A and HNPP. STR�PCR methods detect
three different alleles in CMT1A duplication in
combination with semi�quantitative dosage meas�
urement. Because of its advantages in cost,
amount of DNA sample required, labor, and turn�
around time, the STR�PCR method has been
widely used for differential molecular diagnosis of
CMT1A/HNPP polyneuropathy.
Using this method in our investigation the pres�
ence or absence of 1.5 Mb CMT1A duplication or
HNPP deletion was determined by allelic variants
analysis of four STR�loci (D17S122, D17S921,
D17S1358 and D17S2226) localized in the dupli�
cation/deletion region. Previously we provide the
population study of given STRs in control Ukrainian
population group [13]. The distribution of genotypes
of all loci was in agreement with the expected val�
ues of the exact Hardy�Weinberg equilibrium. It
has been also showed the random allele association
at significance level (p < 0.05) for every pair of loci
in Ukrainian population. However, due to limita�
tions in sensitivity (not direct gene dosage analy�
sis) the present STR�test system informativity for
CMT1A/HNPP analysis using PCR�STR method
in our population was calculated to be nearly 95 %.
So for current analysis of 17p11.2 rearrange�
ments by STR�PCR method we selected the CMT
families with autosome dominant or unclear inheri�
tance (cases without family history of CMT) –
57 families. The STR alleles discrimination had
been carried out by ALF�express fragment analysis
(fig. 2, a). CMT1A�duplication has been identified
in 19 families with clinical CMT1 symptoms and
autosome dominant inheritance with the excep�
ISSN 0564–3783. Цитология и генетика. 2009. № 138
N.V. Hryshechenko, L.A. Livshits
Fig. 3. Case with de novo CMT1A�duplication: a –
D17S2226 locus; 1 – mother (without CMT1A duplication),
genotypes – 4/5; 2, 3 – monozygotic twins (have CMT1A
duplication), genotypes – 2/4/5; 4 – father (without
CMT1A duplication), genotypes – 2/5. ALF�express fluoro�
gramm, 6 % PAAG; b – pedigree of the CMT1A�family
with de novo mutation
ІSSN 0564–3783. Цитология и генетика. 2009. № 1 39
Analysis of 17p11.2 chromosome region rearrangements in CMT1 patients from Ukraine
Fig. 4. Amplification plots of peripheral myelin protein 22 (PMP22) and albumin (ALB), all in duplicates: of: a – normal
control; b – CMT1A patient with PMP22; c – HNPP patient with PMP22
tion of two families. In these two families we sup�
posed the 17p11.2�duplication in probands occurred
due to mutation in germ cells in one of probands’
parents (de novo mutation). This assumption has
been made according to this mutation had not
been identified in anyone of the parents (Fig. 3). In
these families the paternity was confirmed by addi�
tional STR loci analysis. To adduce evidence of de
novo mutation in these families additional quanti�
tative dosage analysis has been done.
In one family HNPP�deletion have been iden�
tified (fig. 2, b). The patients (father and son) had
specific HNPP clinical symptoms – recurrent
neuralgic amyotrophy with sensory loss related to
nerve palsies. The son has paternal chromosome
with deletion (only maternal origin alleles have
been revealed in every of investigated STR�loci).
In this family the paternity was confirmed by addi�
tional 9Y�STR loci analysis (data not shown).
So STR�PCT method revealed to be useful for
prior CMT1A/HNPP detection. But in cases with
de novo mutation as well as in 2 non�informative
cases (the cases with unclear dosage allele inter�
pretation, because of probands were heterozygous
only by 1 STR locus) additional method of analysis
had to be used. In all questionable cases including
1 family with HNPP�deletion direct quantitative
PMP22 gene dosage analysis has been provided. As
a control standards DNA samples with previously
PMP22 copy number identified by STR�PCR
method were used (5 samples with 3 PMP22 copies
and 6 samples with 2 PMP22 copies).
For accurate PMP22 gene dosage detection in
this study we used real�time quantitative PCR with
SYBR Green I dye, as it was described previously
[11] with our own modifications.
The PCR amplification plots of a normal con�
trol sample (Fig. 4, a) showed nearly identical Ct
value of PMP22 triplicates, compared with that of
albumin. In samples positive for the PMP22 dupli�
cation (CMT1A), the Ct value of PMP22 (Ct
Mean PMP22 = 23,91) showed a decrease of
about – 0,85, compared with that of albumin (Ct
Mean ALB = 24,76) (Fig. 4, b), whereas, in sam�
ples positive for the PMP22 deletion (HNPP), the
Ct value of PMP22 (Ct Mean PMP22 = 25,86)
showed an increase of about 0,97, compared with
that of albumin (Ct Mean ALB = 24,89) (Fig. 4,
c). The ��Ct ratio means in normal controls,
including asymptomatic normal family members,
were 0,93–1,30. The ��Ct ratio means in
CMT1A patients were 1,50–2,04. The ��Ct ratio
means in 2 HNPP patients were 0,54 and 0,57. No
overlap was observed between CMT1A or HNPP
patients and normal controls.
Results of real�time quantitative PCR analysis
results confirmed the STR�PCR analysis in des�
cribed HNPP�family as well as the absence of
CMT1A duplication in previously non�informative
samples. We also verified our hypothesis that in two
families under investigation 17p11.2�dupliation in
probands appeared as de novo mutation (Table). In
both families each of parents had 2 PMP22 gene
copies (��Ct ratios were in range 1,1–1,3),
whereas a sick a girl from one family and two twins
from the other had 3 copies of PMP22 gene
(��Ct ratios were in range 1,64; 1,65; 1,50 corre�
spondingly).
So it has been shown that detection of the
PMP22 duplication and deletion by real�time
quantitative PCR is fast and sensitive and lets us
fulfil direct quantitative gene dosage analysis. The
method takes only 2 hr and does not require post�
PCR processing. In addition, DNA samples of
parents are not required to determine 17p11.2
rearrangements in proband. This advantage is the
most useful in HNPP�cases if only proband’s
DNA is available for 17p11.2�deletion analysis.
Conclusions. The methods of 17p11.2 duplica�
tion/deletion identification have been elaborated:
STR allelic variants analysis and direct PMP22 gene
dosage measuring by means of quantitative Real�
Time PCR. It has been shown that Real�Time PCR
using SYBR Green I dye is highly sensitive, specif�
ISSN 0564–3783. Цитология и генетика. 2009. № 140
N.V. Hryshechenko, L.A. Livshits
Real�time data in families with de novo mutations
in probands
Identifier
PMP22 ALB
��Ct ratio
Ct Mean
Proband’s mother
Proband’s father
Proband
23,66
23,60
22,68
23,97
23,73
23,40
1,24
1,09
1,64
Family 1
Family 2
Proband’s mother
Proband’s father
Proband 1
Proband 2
25,23
24,42
24,14
24,02
25,37
24,63
24,87
24,61
1,10
1,15
1,65
1,50
ic, and reproducible in detecting PMP22 duplica�
tion and deletion in patients with CMT1A and
HNPP. We also indicate high level of de novo cases
with 17p11.2�duplication – in 2 of 19 CMT1A�
families. It has been shown the 17p11.2 chromo�
some region duplication/deletion association with
CMT1A and HNPP clinical phenotypes which
may be used in differential diagnosis of this type of
CMT polyneuropathy.
Н.В. Грищенко, Л.А. Лившиц
АНАЛИЗ ХРОМОСОМНЫХ ПЕРЕСТРОЕК
В ОБЛАСТИ 17р11.2 У БОЛЬНЫХ ШМТ1
ИЗ УКРАИНЫ
Проведена детекция и анализ хромосомных пере�
строек области 17p11.2 у пациентов с болезнью Шарко�
Мари�Тус (ШМТ) из Украины. Разработаны два взаи�
модополняющих метода идентификации дуплика�
ций/делеций в области 17p11.2: анализ аллельных ва�
риантов STR�локусов и прямой анализ дозы гена
РМР22 с использованием ПЦР в реальном времени.
Выявлен высокий уровень de novo дупликаций в ука�
занном хромосомном регионе. Показана ассоциация
дупликаций/делеций в области 17p11.2 с клинически�
ми фенотипами ШМТ1А и HNPP, что может быть ис�
пользовано для дифференциальной диагностики
упомянутых типов полиневропатии ШМТ.
Н.В. Грищенко, Л.А. Лівшиць
АНАЛІЗ ХРОМОСОМНИХ ПЕРЕБУДОВ
У ДІЛЯНЦІ 17р11.2 У ХВОРИХ ШМТ1
З УКРАЇНИ
Проведено детекцію та аналіз хромосомних пере�
будов ділянки 17p11.2 у пацієнтів з хворобою Шарко�
Марі�Тус (ШМТ) з України. Розроблено два взаємо�
доповнюючі методи ідентифікації дуплікацій/делецій
у ділянці 17p11.2: аналіз алельних варіантів STR�локу�
сів та прямий аналіз дози гена РМР22 із використан�
ням ПЛР у реальному часі. Виявлено високий рівень
de novo дуплікацій цієї хромосомної ділянки. Показа�
но ассоціацію дуплікацій/делецій у ділянці 17p11.2
із клінічними фенотипами ШМТ1А та HNPP, що мо�
же бути використано для диференційної діагностики
даних типів поліневропатії ШМТ.
REFERENCES
1. Skre H. Genetic and clinical aspects of Charcot�Marie�
Tooth’s disease // Clin. Genet. – 1974. – 6, № 2. –
P. 98–118.
2. Bouche P. How to interpret the electromyogram (in
neuromuscular diseases) // Rev. Prat. 1983. – 33, №
11. – P. 549–555.
3. Boerkoel C.F., Takashima H., Garcia C.A. et al. Char�
cot�Marie�Tooth disease and related neuropathies:
mutation distribution and genotype�phenotype corre�
lation // Ann. Neurol. – 2002. – 51. № 2. – P. 190–
201.
4. Chance P.F., Alderson M.K., Leppig K.A. et al. DNA
deletion associated with hereditary neuropathy with
liability to pressure palsies // Cell. – 1993. – 72, № 1. –
P. 143–51.
5. Pentao L., Wise C.A., Chinault A.C., Patel P.1., Lupski
J.R. Charcot�Marie�Tooth type IA duplication appears
to arise from recombination at repeat sequences flank�
ing the 1.5 Mb monomer unit // Nature Genet. – 1992. –
2. – P. 292–300.
6. Lupski JR. Charcot�Marie�Tooth disease: lessons in
genetic mechanisms // Mol. Med. – 1998. – 4, № 1. –
P. 3–11.
7. Badano J.L., Inoue K., Katsanis N., Lupski J.R. New
polymorphic short tandem repeats for PCR�based
Charcot�Marie�Tooth disease type 1A duplication
diagnosis // Clin. Chem. – 2001. – 47, №5. – P. 838–
843.
8. Kashork C.D., Lupski J.R., Shaffer L.G. Prenatal diag�
nosis of Charcot�Marie�Tooth disease type 1A by inter�
phase fluorescence in situ hybridization // Prenat
Diagn. – 1999. – 19. – P. 446–449.
9. Hensels G.W., Janssen E., Hoogendijk J.E. et al.
Quantitative measurement of duplicated DNA as a
diagnostic test for Charcot�Marie�Tooth disease type la //
Chin. Chem. – 1993. – 39. – P. 1845–1849.
10. Rowland J.S., Barton D.E., Taylor G.R. et al. A compar�
ison of methods for gene dosage analysis in HMSN
type 1 // Med. Genet. – 2001. – 38. – P. 90–95.
11. Sang�Wun Kim, Kwang�Soo Lee, Hyun�Seok Jin et al.
Rapid detection of duplication/deletion of the
PMP22 gene in patients with Charcot�Marie�Tooth
disease type 1A and hereditary neuropathy with liabili�
ty to pressure palsy by real�time quantitative PCR using
SYBR green I dye // J. Korean Med. Sci. – 2003. – 18. –
P. 727–732.
12. Maniatis T., Fritsch E.F., Sambrook J. Molecular
cloning : a laboratory manual. – Cold Spring Harbor
Laboratory, 1982. – 545 p.
13. Grishchenko N.V., Bychkova A.M., Pichkur N.A et al.
Study of the duplication of 17Р11.2–12 chromosome
region for the patients with hereditary motor and sen�
sory neuropathy type 1A // Cytol. and Genet. – 2003. –
37, № 6. – Р. 55–59.
14. Hryshchenko N.V., Kravchenko S.A., Livshits L.A.
Polymorphic short tandem repeats for PCR�based
diagnosis of the Charcot�Marie�Tooth 1A duplication
in Ukraine // Cytol. and Genet. – 2005. – 39, № 5. –
P. 56–61.
Received 11.02.08
ІSSN 0564–3783. Цитология и генетика. 2009. № 1 41
Analysis of 17p11.2 chromosome region rearrangements in CMT1 patients from Ukraine
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