Demonstration of Two alpha-Globin Genes per Human Haploid Genome for Normals and Hb J Mexico

Tolstoshev, Paul; Eskdale, Joyce; Williamson, Robert; Verdier, Gérard; Godet, Jacqueline; Nigon, Victor; Trabuchet, G; Benabadji, M.

doi:10.1111/j.1432-1033.1977.tb11725.x

Cited by 18 publications

(6 citation statements)

References 20 publications

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“…However, the «n light-chain mRNA hybridized with a Cr(|/2 value of 1.3 X 10~3 (Figure 6A) which was significantly lower than expected. Assuming a combined size for the « and f) human globin mRNAs of about 1400 nucleotides (Proudfoot et al, 1977;Tolstoshev et al, 1977) and taking 1200 nucleotides (Figure I D) for the ki 1 mRNA, the expected C\t 1/2 value for a ki 1 mRNA preparation that is 75% pure would be 1.8 X 10-3. These results demonstrate that the sole use of globin mRNA as a complexity standard in our case would have resulted in an overestimation of mRNA purity.…”

Section: Resultsmentioning

confidence: 99%

Isolation, purification, and properties of mouse heavy-chain immunoglobulin mRNAs

Marcu¹,

Valbuena²,

Perry³

1978

Biochemistry

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A procedure is described for the isolation of highly purified heavy-chain immunoglobulin mRNAs from a variety of mouse plasmacytomas (IgA, IgG, and IgM producers). The use of fresh tissue and the rapid isolation and direct extraction of membrane-bound polyribosomes were found to be essential in obtaining large quantities of undegraded heavy-chain mRNAs. The individual mRNAs were purified by two cycles of oligo(dT)-cellulose chromatography, sodium dodecyl sulfate--sucrose gradient centrifugation, and electrophoresis on 98% formamide containing polyacrylamide gels. When added to a cell-free protein-synthesizing system from wheat germ, the MPC-11 gamma2b and H2020 alpha heavy-chain mRNAs efficiently directed the synthesis of a predominant product of 55 000 molecular weight, while the synthesis of a 70 000 dalton protein in addition to other lower molecular weight polypeptides were observed with MOPC 3741 mu mRNA. All of these proteins were immunoprecipitable with class-specific heavy-chain antisera, and in the case of the gamma2b in vitro products good correspondence in a comparative trypsin--chymotrypsin fingerpring with in vivo labeled gamma2b heavy chain was observed. The gamma2b and a alpha heavy-chain mRNAs possessed a chain length of approximately 1800 nucleotides and the mu mRNA a size of approximately 2150 nucleotides when examined under stringent denaturation conditions. The purities of the alpha, gamma2b, and mu mRNAs were estimated to be 60--80%, 50--70%, and 50--83%, respectively, on the basis of their hybridization rates with cDNA probes in comparison to mRNA standards of known complexity. Heavy-chain mRNAs of the same class isolated from different mouse strains (Balb/C or NZB) display no detectable sequence differences in cross hybridization experiments, even though the cDNA--mRNA hybrids are submitted to stringent S1 nuclease digestion. These results indicate that allotypic determinants represent only a minor fraction of the heavy-chain constant region sequence in the mouse.

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Section: Resultsmentioning

confidence: 99%

Isolation, purification, and properties of mouse heavy-chain immunoglobulin mRNAs

Marcu¹,

Valbuena²,

Perry³

1978

Biochemistry

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“…The relative output of the normal non-a-globin genes (G7, A), 6, and f3) does not correspond directly to the number of loci, suggesting that mechanisms other than simple gene dosage are important. However, even the percentages of an a-chain variant may not always correlate with the number of gene loci present (38).…”

Section: Discussionmentioning

confidence: 99%

Linkage of alpha G-Philadelphia to alpha-thalassemia in African-Americans .

Surrey¹,

Ohene‐Frempong²,

Rappaport³

et al. 1980

Proc. Natl. Acad. Sci. U.S.A.

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ABSIRACT We have studied the inheritance of the a-chain hemoglobin variant Hb G-Philadelphia (a268AsILys#2) in two African-American families. Expression of the a-globin loci was monitored by the percentage of Hb G in these individuals. The variant represented approximately 33% of the total adult hemoglobin in some an 50% in others. a-Globin gene fragments were analyzed by using restriction endonucleases that cleave outside (EcoRI), within (HindIII), and between (Bgl II) the normal duplicated a-globin loci (aa/aa). Individuals having 33% variant lack one functioning a gene (aG/aa); those with 50% variant lack two genes, one missing on each chromosome (aG/a). Inheritance of aG was therefore linked to that of a chromosome with only one functional a-globin gene locus. This locus is probably the result of a nonhomologous crossover. Our results a so sugest equal expression of the a-globin loci in humans because the percentages of the variant could be explained solely on the basis of the total number of a genes present. The percentages of Hb G as well as other hematologic data all were consistent with the number of a-globin genes identified by restriction endonuclease mapping Gene mapping yields a more precise determination of the number of a-globin genes than does study of globin synthesis.The existence of two a-globin genes per haploid genome in humans is now well established (1, 2). Solution hybridization (3-6) and restriction endonuclease analyses of DNA (7-11) have shown that the most common cause of a-thalassemia is deletion of structural gene material, although nondeletion types also have been reported (12, 13). The four classical disorders due to a-thalassemia (silent carrier, a-thalassemia trait, Hb H disease, and hydrqps fetalis) are most frequently caused by deletion of one to four a-globin genes.In Asians, deletion of two a-globin genes on the same chromosome or of only one gene is common, whereas in Africans, deletion of only a single a-globin locus per chromosome is the predominant form (9, 10). The a-thalassemia syndromes in Asians commonly arise by gene deletion of either the 5' a-globin locus or both 5' and 3' a-globin loci; in Africans and Mediterraneans, as well as some Asians, generation of a single a-globin locus on a chromosome is due to a nonhomologous crossover (12,14).The most common a-chain variant in African-Americans is aCGPhiladelphia (aX68.Asn-bsy; aG), which has been reported to represent 20%, 30%, or 40% of the total.number of a-chains present (15). It has been suggested that this distribution may be due to the linkage of the variant a-globin gene to a-thalassemia (16, 17).We have investigated the expression of a-thalassemia in two African-American families with aG by using hematologic data, globin synthesis, and analyses of a-globin genes in restriction endonuclease digests of DNA. MATERIALS AND METHODSHematologic Studies. Nine individuals from two AfricanAmerican families were studied. In addition to aG, family 2 also had Hb S and f+-thalassemia genotypes (see Table 1). Hematologic stu...

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“…They were hybridized in cDNA excess to complementary DNA specific for a-globin mRNA [20], The extent of hybridization of cellular DNA to cDNAa was determined, after Sinuclease digestion, exactly as already reported [21].…”

Section: Methodsmentioning

confidence: 99%

Hemoglobin H Disease from Algeria: Genetic and Molecular Characterization

Tabone¹,

Henni²,

Belhani³

et al. 1981

Acta Haematol

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A case of Hb H disease from Algeria was studied at the genetic and molecular level in order to delineate the pattern of α-thalassemia in the Mediterranean population. The family study indicated that both parents had the hematological and clinical manifestation of α-thalassemia trait and that the affected sibling had homozygous α-thalassemia with 5.6% Hb H, microcytosis and an α-/non-α-biosynthetic ratio of 0.64. Hybridization in globin cDNAα excess suggested that the molecular defect responsible for this form of α-thalassemia is a partial deletion of the haploid stock of α-globin genes. The Algerian case of Hb H disease studied thus differs from Asian and Negro cases by the mode of inheritance of the α-thalassemia mutation involved.

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Demonstration of Two alpha-Globin Genes per Human Haploid Genome for Normals and Hb J Mexico

Cited by 18 publications

References 20 publications

Isolation, purification, and properties of mouse heavy-chain immunoglobulin mRNAs

Isolation, purification, and properties of mouse heavy-chain immunoglobulin mRNAs

Linkage of alpha G-Philadelphia to alpha-thalassemia in African-Americans .

Hemoglobin H Disease from Algeria: Genetic and Molecular Characterization

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