Rapid detection of point mutations and polymorphisms of the α-globin genes by DGGE and SSCA

Harteveld, Kees L.; Heister, Angelien J.G.A.M.; Giordano, Piero C.; Losekoot, Monique; Bernini, L. F.

doi:10.1002/(sici)1098-1004(1996)7:2<114::aid-humu5>3.0.co;2-c

Cited by 61 publications

(16 citation statements)

References 25 publications

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“…Hematological indices were automatically measured on a Coulter Counter ABX Micros 60 (Helena Laboratories, Beaumont, TX, USA). The a-globin genes were amplified using specific primers (M13S13 and S6 for a2 and M13S13 and S8 for a1), and sequenced by internal primers as previously described (17). The Hb H and Hb A 2 levels were measured densitometrically.…”

Section: Methodsmentioning

confidence: 99%

Molecular Basis of Hb H Disease in Southwest Iran

Yavarian¹,

Karimi²,

Zorai³

et al. 2005

LHEM

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Although alpha0-thalassemia (thal) defects are not very frequent in the Iranian population, Hb H disease does occur in the country. We have analyzed the alpha gene cluster of 13 patients showing the presence of Hb H to establish the molecular background of this disease in southwest Iran (Shiraz and Hormozgan provinces). Using gap-polymerase chain reaction (gap-PCR) and direct DNA sequencing we have found the --MED-I deletion, the polyadenylation signal (poly A) mutations alphaT-Saudi alpha and alphaT-Turkish alpha and Hb Constant Spring (Hb CS) in association with the common -alpha3.7 deletion. This study has revealed that: 1) at least six genotypes are responsible for Hb H disease in the area: .-alpha3.7/ --MED-I; -alpha3.7/alphaT-Saudi alpha; alphaT-Saudi alpha/alphaT-Saudi alpha; alphaCSalpha/--MED-I; --MED-I/alphaT-Turkish alpha; and the atypical forms of Hb H disease -alpha3.7/alphaCSalpha. 2) The molecular background of Hb H disease in the southwest area of Iran is more similar to the Mediterranean type than to the Southeast Asian. 3) Hb Bart's hydrops fetalis syndrome and mild, intermediate or severe postnatal Hb H disease conditions can be expected, but at a relatively low incidence. 4) The diagnostic flowchart for patients with microcytic hypochromic anemia should include iron deficiency, beta-thal, alpha+- and alpha0-thal analyses.

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Section: Methodsmentioning

confidence: 99%

Molecular Basis of Hb H Disease in Southwest Iran

Yavarian¹,

Karimi²,

Zorai³

et al. 2005

LHEM

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“…Analysis of point mutations in the a-globin genes. The a2 and a1 genes were amplified as described earlier (Harteveld et al, 1996) and the amplified products were subjected to DNA sequencing using the Big Dye terminator kit (Applied Biosystems, Foster City, CA, USA) on the ABI 310 genetic analyzer (Applied Biosystems). Fig 1. (A) Schematic representation of a-globin gene cluster indicating the extent of the ) ) SA deletion and its neighbouring deletions.…”

Section: Methodsmentioning

confidence: 99%

Determination of the breakpoint and molecular diagnosis of a common α‐thalassaemia‐1 deletion in the Indian population

Shaji¹,

Eunice²,

Baidya

et al. 2003

Br J Haematol

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Summary. The previously described South African type a-thalassaemia-1 mutation was identified in Indian HbH patients using a polymerase chain reaction (PCR) strategy. A multiplex PCR assay was devised to detect heterozygotes and homozygotes. This a-thalassaemia-1 mutation was found to be the commonest determinant causing HbH disease in this population. In one family this mutation was found in combination with a novel splice donor mutation a2 IVS I-1 (G fi A). Characterization of the breakpoint junction sequence revealed, in addition to a 23 kb deletion, that there was an addition of 160 bp bridging the breakpoints. Similar to other deletions in the a-globin gene cluster, there is an Alu repeat-mediated mechanism for the origin of the deletion. Keywords: a-thalassaemia, PCR, India, AluThe a-thalassaemias are genetic defects characterized by the decrease or complete suppression of synthesis of the a-globin polypeptide chains of haemoglobin. They are the most common single-gene diseases in the world. The a-globin genes are duplicated (a1 and a2) and located within the a gene cluster f2)wf1)wa2)wa1)a2)a1)h1 on the distal short arm region of chromosome 16 (Higgs et al, 1989). This cluster contains four genes and three pseudogenes. Deletions of one ()a) or both () ) )) of these cis-linked genes are the most common causes of the a-thalassaemias. Around 25 different two-gene cis deletions () )) and eight single-gene deletions ()a)

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“…Known Mediterranean nondeletional α‐thalassaemia mutations in the α2‐globin genes were investigated as previously described (Hall et al , 1993; Traeger‐Synodinos et al , 1993). Further characterization of α‐globin gene point mutations was carried out by DGGE analysis (Harteveld et al , 1996).…”

Section: Methodsmentioning

confidence: 99%

Molecular, haematological and clinical studies of the −101 C → T substitution of the β‐globin gene promoter in 25 β‐thalassaemia intermedia patients and 45 heterozygotes

et al. 1999

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Summary. We report the clinical, haematological, biosynthetic and molecular data of 25 double heterozygote b-thalassaemia intermedia patients and 45 b-thalassaemia heterozygotes with the C ! T substitution at nucleotide position À101 from the Cap site, in the distal CACCC box of the b-globin gene promoter. This mutation is considered the most common amongst the silent b-thalassaemia mutations in Mediterranean populations. Of the 25 compound heterozygotes for the b À101 C ! T and common severe bthalassaemia mutations, all but one had mild thalassaemia intermedia preserving haemoglobin levels around 9´5 g/dl and haemoglobin F levels < 25%. The only transfused patient was characterized to have an additional a-globin gene.Strict assessment of haematological and biosynthetic ®nd-ings in the heterozygotes for the b À101 C ! T mutation (excluding six cases with an a-globin gene defect) demonstrated that less than half of them had completely normal (silent) haematology; the remainder had either high haemoglobin A 2 values (in the range of 3´7±5´1%) and/or low red cells indices and/or raised haemoglobin F values. The a/non-aglobin chain synthesis ratios were generally raised, with mean 1´44 (1´07±2´10). Amongst the parents of the compound heterozygotes, who were not selected for molecular analysis following haematological screening, half of the cases were completely silent.Interaction with severe b-thalassaemia mutations always resulted in the clinical phenotype of mild non-transfusiondependent thalassaemia intermedia.

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Rapid detection of point mutations and polymorphisms of the α-globin genes by DGGE and SSCA

Cited by 61 publications

References 25 publications

Molecular Basis of Hb H Disease in Southwest Iran

Molecular Basis of Hb H Disease in Southwest Iran

Determination of the breakpoint and molecular diagnosis of a common α‐thalassaemia‐1 deletion in the Indian population

Molecular, haematological and clinical studies of the −101 C → T substitution of the β‐globin gene promoter in 25 β‐thalassaemia intermedia patients and 45 heterozygotes

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