1.A family study of HbS in a Malay family by molecular analysis.
Hafiza, Alauddin ; Noor, Hamidah Hussin ; Noor, Farisah A Razak ; Azlin, Ithnin ; Ainoon, Othman
The Malaysian Journal of Pathology 2010;32(2):137-41
Sickle cell disease (SCD) is an inherited red cell disorder, characterized by the tendency of haemoglobin S or sickle haemoglobin to polymerize and assume a characteristic sickle shape. Molecular analysis has been the mainstay of detection method when confirmation is required. Previously a polymerase chain reaction (PCR)-based restriction enzyme analysis was used for this purpose. A simple bidirectional allele-specific amplification, recently described by Waterfall in 2001 was used to detect the GAG --> GTG mutation on codon 6 of the beta globin gene. Two sets of primers for the mutant and the wild type alleles were used in a single PCR reaction to amplify the regions of interest. The resultant PCR products will produce two fragments at 517 and 267 base pair (bp) respectively. This report highlights the investigations for SCD in the family of a 16-year old girl with recurrent painful crisis affecting the lower limbs whereby the family members are asymptomatic for the disease. Her haemoglobin electrophoresis at an alkaline pH showed dense bands at the HbS and HbF regions, while her father and two sisters had bands at HbS, HbF and HbA. The PCR analysis showed that she was homozygous for the mutation by the presence of only one band at 267 bp fragment, while the father and her sisters were heterozygotes, with the presence of two bands at 267 as well as 517 bp fragments. DNA sequencing of the sample confirmed the mutation. In conclusion, this case report highlighted the simple and cheap yet practical method for molecular confirmation of the presence of HbS gene in subjects with homozygous or heterozygous state of the condition.
Anemia, Sickle Cell/*diagnosis
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Anemia, Sickle Cell/*genetics
;
Base Sequence
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Fathers
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Hemoglobin, Sickle/*genetics
;
Heterozygote
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Homozygote
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Malaysia
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Mutation
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Nucleic Acid Amplification Techniques
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Pedigree
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Polymerase Chain Reaction
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Siblings
2.Molecular characteristic of alpha thalassaemia among patients diagnosed in UKM Medical Centre
Raja Zahratul AZMA ; AINOON Othman ; HAFIZA Alauddin ; AZLIN Ithnin ; Noor FARISAH Abdul Razak ; Nor HIDAYATI Sardi ; Noor HAMIDAH Hussin
The Malaysian Journal of Pathology 2014;36(1):27-32
Alpha (α) thalassaemia is the most common inherited disorder in Malaysia. The clinical severity
is dependant on the number of α genes involved. Full blood count (FBC) and haemoglobin (Hb)
analysis using either gel electrophoresis, high performance liquid chromatography (HPLC) or
capillary zone electrophoresis (CE) are unable to detect definitively alpha thalassaemia carriers.
Definitive diagnosis of α-thalassaemias requires molecular analysis and methods of detecting
both common deletional and non-deletional molecular abnormailities are easily performed in any
laboratory involved in molecular diagnostics. We carried out a retrospective analysis of 1623 cases
referred to our laboratory in Universiti Kebangsaan Malaysia Medical Centre (UKMMC) for the
diagnosis of α-thalassaemia during the period October 2001 to December 2012. We examined the
frequency of different types of alpha gene abnormalities and their haematologic features. Molecular
diagnosis was made using a combination of multiplex polymerase reaction (PCR) and real time
PCR to detect deletional and non-deletional alpha genes relevant to southeast Asian population.
Genetic analysis confirmed the diagnosis of α-thalassaemias in 736 cases. Majority of the cases
were Chinese (53.1%) followed by Malays (44.2%), and Indians (2.7%). The most common gene
abnormality was αα/--SEA (64.0%) followed by αα/-α3.7 (19.8%), -α3.7 /--SEA (6.9%), αα/ααCS (3.0%),
--SEA/--SEA (1.2%), -α3.7/-α3.7 (1.1%), αα/-α4.2 (0.7%), -α4.2/--SEA (0.7%), -α3.7/-α4.2 (0.5%), ααCS/--
SEA (0.4%), ααCS/ααCd59 (0.4%), ααCS/ααCS (0.4%), -α3.7/ααCd59 (0.3%), αα/ααCd59 (0.1%), αα Cd59/
ααIVS I-1 (0.1%), -α3.7/ααCS (0.1%) and --SEA /ααCd59 (0.1%). This data indicates that the molecular
abnormalities of α-thalassaemia in the Malaysian population is heterogenous. Although α-gene
deletion is the most common cause, non-deletional α-gene abnormalities are not uncommon and at
least 3 different mutations exist. Establishment of rapid and easy molecular techniques is important
for definitive diagnosis of alpha thalassaemia, an important prerequisite for genetic counselling to
prevent its deleterious complications.
Thalassemia
;
Patients
3.Co-inheritance of compound heterozygous Hb Constant Spring and a single –α3.7 gene deletion with heterozygous δβ thalassaemia: A diagnostic challenge
Raja Zahratul Azma ; Ainoon Othman ; Norazlina Azman ; Hafiza Alauddin ; Azlin Ithnin ; Nurasyikin Yusof ; Noor Farisah Razak ; Nor Hidayati Sardi ; Noor Hamidah Hussin
The Malaysian Journal of Pathology 2012;34(1):57-62
Haemoglobin Constant Spring (Hb CS) mutation and single gene deletions are common underlying
genetic abnormalities for alpha thalassaemias. Co-inheritance of deletional and non-deletional alpha
(α) thalassaemias may result in various thalassaemia syndromes. Concomitant co-inheritance with
beta (β) and delta (δ) gene abnormalities would result in improved clinical phenotype. We report here
a 33-year-old male patient who was admitted with dengue haemorrhagic fever, with a background
history of Grave’s disease, incidentally noted to have mild hypochromic microcytic red cell indices.
Physical examination revealed no thalassaemic features or hepatosplenomegaly. His full blood
picture showed hypochromic microcytic red cells with normal haemoglobin (Hb) level. Quantitation
of Hb using high performance liquid chromatography (HPLC) and capillary electrophoresis (CE)
revealed raised Hb F, normal Hb A2 and Hb A levels. There was also small peak of Hb CS noted in
CE. H inclusions was negative. Kleihauer test was positive with heterocellular distribution of Hb
F among the red cells. DNA analysis for α globin gene mutations showed a single -α-3.7 deletion
and Hb CS mutation. These fi ndings were suggestive of compound heterozygosity of Hb CS and
a single -α-3.7 deletion with a concomitant heterozygous δβ thalassaemia. Co-inheritance of Hb
CS and a single -α-3.7 deletion is expected to result at the very least in a clinical phenotype similar
to that of two alpha genes deletion. However we demonstrate here a phenotypic modifi cation of α
thalassemia presumptively as a result of co-inheritance with δβ chain abnormality as suggested by
the high Hb F level.